Pyrazine derivatives as enac blockers

ABSTRACT

Compounds of Formula I: 
     
       
         
         
             
             
         
       
     
     and pharmaceutically acceptable salts and solvates thereof, wherein R 1 , R 2 , R 3 , R 4 , R 5 , R 6 , R 7 , R 8 , R 9 , and R 10  have the meanings as indicated in the specification, are useful for treating diseases mediated by blockade of the epithelial sodium channel. Pharmaceutical compositions that contain the compounds and processes for preparing the compounds are also described.

RELATED APPLICATIONS

This application is a Continuation of U.S. application Ser. No.13/232,175, filed Sep. 14, 2011, which claims the benefit of U.S.Provisional Application No. 61/383,985, filed on Sep. 17, 2010, and U.S.Provisional Application No. 61/524,495, filed on Aug. 17, 2011, theentire teachings of which are incorporated by reference.

This invention relates to organic compounds and their use aspharmaceuticals, in particular for the treatment of inflammatory,obstructive or allergic diseases and conditions, particularly aninflammatory or obstructive airways disease or mucosal hydration.

In one aspect, the invention provides a compound of Formula I:

or a pharmaceutically acceptable salt or solvate thereof, whereinR¹ is H, halogen, C₁-C₈-alkyl, C₁-C₈-haloalkyl, C₁-C₈-haloalkoxy,C₃-C₁₅-carbocyclic group, nitro, cyano, a C₆-C₁₅-membered aromaticcarbocyclic group, or a C₁-C₈-alkyl substituted by a C₆-C₁₅-memberedaromatic carbocyclic group;R², R³, R⁴ and R⁵ are each independently selected from H and C₁-C₆alkyl;R⁶, R⁷, R⁸ and R⁹ are each independently selected from H; SO₂R¹⁶; aryloptionally substituted by one or more Z groups; a C₃-C₁₀ carbocyclicgroup optionally substituted by one or more Z groups; C₃-C₁₄heterocyclic group optionally substituted by one or more Z groups; C₁-C₈alkyl optionally substituted by an aryl group which is optionallysubstituted by one or more Z groups, a C₃-C₁₀ carbocyclic groupoptionally substituted by one or more Z groups or a C₃-C₁₄ heterocyclicgroup optionally substituted by one or more Z groups;R¹⁰ is represented by the formula 2:

—(C₀-C₃alkylene)-B—X—(CR^(11a)R^(12a))_(m)—(CR^(11b)R^(12b))_(n)—(CR^(11c)R^(12c))_(p)—C(O)OR¹³,

wherein the alkylene groups are optionally substituted by one or more Zgroups;B is aryl optionally substituted by one or more Z groups;X is selected from a bond, —NR¹⁵(SO₂)—, —(SO₂)NR¹⁵—, —(SO₂)—,—NR¹⁵C(O)—, —C(O)NR¹⁵—, —NR¹⁵C(O)NR¹⁷—, —NR¹⁵C(O)O—, —NR¹⁵—, C(O)O,OC(O), C(O), O and S;R^(11a), R^(11b), R^(11c), R^(12a), R^(12b) and R^(12c) are eachindependently selected from H and C₁-C₆ alkyl; orR^(11a) and R^(12a) together with the carbon atom to which they areattached form a 3- to 8-membered cycloalkyl group; orR^(11b) and R^(12b) together with the carbon atom to which they areattached form a 3- to 8-membered cycloalkyl group; orR^(11c) and R^(12c) together with the carbon atom to which they areattached form a 3- to 8-membered cycloalkyl group;R¹³ is selected from (C₁-C₃ alkyl)-C(O)NR²²R²³; (C₁-C₃ alkyl)-C(O)OR²³;and (C₁-C₃ alkyl)-NR²³C(O)R²²R¹⁵ and R¹⁷ are each independently selected from H and C₁-C₆ alkyl;R¹⁶ is selected from C₁-C₈ alkyl, aryl and a 3- to 14-memberedheterocyclic group, the heterocyclic group including one or moreheteroatoms selected from N, O and S;Z is independently selected from —OH, aryl, —O-aryl, C₇-C₁₄ aralkyl,—O—C₇-C₁₄ aralkyl, C₁-C₆ alkyl, C₁-C₆ alkoxy, —NR¹⁹(SO₂)R²¹,—(SO₂)NR¹⁹R²¹, —(SO₂)R²⁰, —NR¹⁹C(O)R²⁰, —C(O)NR¹⁹R²⁰, —NR¹⁹C(O)NR²⁰R¹⁸,—NR¹⁹C(O)OR²⁰, —NR¹⁹R²¹, C(O)OR¹⁹, —C(O)R¹⁹, SR¹⁹, —OR¹⁹, oxo, CN, NO₂,and halogen, wherein the alkyl, alkoxy, aralkyl and aryl groups are eachoptionally substituted by one or more substituents selected from OH,halogen, C₁-C₄ haloalkyl and C₁-C₄ alkoxy;R¹⁸, R²⁰ and R²² are each independently selected from H and C₁-C₆ alkyl;R¹⁹, R²¹ and R²³ are each independently selected from H; C₁-C₈ alkyl;C₃-C₈ cycloalkyl; C₁-C₄ alkoxy-C₁-C₄ alkyl; (C₀-C₄ alkyl)-aryloptionally substituted by one or more groups selected from C₁-C₆ alkyl,C₁-C₆ alkoxy and halogen; (C₀-C₄ alkyl)-3- to 14-membered heterocyclicgroup, the heterocyclic group including one or more heteroatoms selectedfrom N, O and S, optionally substituted by one or more groups selectedfrom halogen, oxo, C₁-C₆ alkyl and —C(O)C₁-C₆ alkyl; —(C₀-C₄alkyl)-O-aryl optionally substituted by one or more groups selected fromC₁-C₆ alkyl, C₁-C₆ alkoxy and halogen; and (C₀-C₄ alkyl)-O-3- to14-membered heterocyclic group, the heterocyclic group including one ormore heteroatoms selected from N, O and S, optionally substituted by oneor more groups selected from halogen, C₁-C₆ alkyl and —C(O)C₁-C₆ alkyl;wherein the alkyl groups are optionally substituted by one or morehalogen atoms, hydroxyl, C₁-C₄ alkoxy, —C(O)NH₂, —C(O)NHC₁-C₆ alkyl or—C(O)N(C₁-C₆ alkyl)₂; orR¹⁹ and R²⁰ together with the nitrogen atom to which they are attachedform a 5- to 10-membered heterocyclic group, the heterocyclic groupincluding one or more further heteroatoms selected from N, O and S, theheterocyclic group being optionally substituted by one or moresubstituents selected from OH; halogen; aryl; 5- to 10-memberedheterocyclic group including one or more heteroatoms selected from N, Oand S; —S(O)₂-aryl; —S(O)₂—C₁-C₆ alkyl; C₁-C₆ alkyl optionallysubstituted by one or more halogen atoms; C₁-C₆ alkoxy optionallysubstituted by one or more OH groups or C₁-C₄ alkoxy; and C(O)OC₁-C₆alkyl, wherein the aryl and heterocyclic substituent groups arethemselves optionally substituted by C₁-C₆ alkyl, C₁-C₆ haloalkyl orC₁-C₆ alkoxy; orR¹⁹ and R²¹ together with the nitrogen atom to which they are attachedform a 5- to 10-membered heterocyclic group, the heterocyclic groupincluding one or more further heteroatoms selected from N, O and S, theheterocyclic group being optionally substituted by one or moresubstituents selected from OH; halogen; aryl; 5- to 10-memberedheterocyclic group including one or more heteroatoms selected from N, Oand S; —S(O)₂-aryl; —S(O)₂—C₁-C₆ alkyl; C₁-C₆ alkyl optionallysubstituted by one or more halogen atoms; C₁-C₆ alkoxy optionallysubstituted by one or more OH groups or C₁-C₄ alkoxy; and —C(O)OC₁-C₆alkyl, wherein the aryl and heterocyclic substituent groups arethemselves optionally substituted by C₁-C₆ alkyl, C₁-C₆ haloalkyl orC₁-C₆ alkoxy; orR¹⁸ and R²⁰ together with the nitrogen atom to which they are attachedform a 5- to 10-membered heterocyclic group, the heterocyclic groupincluding one or more further heteroatoms selected from N, O and S, theheterocyclic group being optionally substituted by one or moresubstituents selected from OH; halogen; aryl; 5- to 10-memberedheterocyclic group including one or more heteroatoms selected from N, Oand S; —S(O)₂-aryl; —S(O)₂—C₁-C₆ alkyl; C₁-C₆ alkyl optionallysubstituted by one or more halogen atoms; C₁-C₆ alkoxy optionallysubstituted by one or more OH groups or C₁-C₄ alkoxy; and —C(O)OC₁-C₆alkyl, wherein the aryl and heterocyclic substituent groups arethemselves optionally substituted by C₁-C₆ alkyl, C₁-C₆ haloalkyl orC₁-C₆ alkoxy; orR²² and R²³ together with the atom(s) to which they are attached form a5- to 10-membered heterocyclic group, the heterocyclic group includingone or more further heteroatoms selected from N, O and S, theheterocyclic group being optionally substituted by one or more Z groups;m is 0, 1, 2 or 3;n is 0, 1, 2 or 3;p is 0, 1, 2 or 3;wherein at least one of m, n or p is not 0.

In another embodiment of the invention as defined anywhere above, R¹ ishalogen. In a further embodiment, R¹ is chlorine

In another embodiment of the invention as defined anywhere above, R²,R³, R⁴ and R⁵ are H.

In another embodiment of the invention as defined anywhere above, R⁶,R⁷, R⁸ and R⁹ are H.

In another embodiment of the invention as defined anywhere above, R¹⁰ is—B—X—(CR^(11a)R^(12a))—(CR^(11b)R^(12b))—C(O)OR¹³.

In a further embodiment, R¹⁰ is—B—(SO₂)NR¹⁵—(CR^(11a)R^(12a))—(CR^(11b)R^(12b))—C(O)OR¹³.

In an alternative yet further embodiment, R¹⁰ is—B—NR¹⁵C(O)NR¹⁷—(CR^(11a)R^(12a))—(CR^(11b)R^(12b))—C(O)OR¹³.

In an alternative embodiment of the invention as defined anywhere above,R¹⁰ is —(C₂alkylene)-B—X—(CR^(11a)R^(12a))—C(O)OR¹³, wherein thealkylene groups are optionally substituted by one or more Z groups. In afurther embodiment, the alkylene groups are unsubstituted. In a yetfurther embodiment, X is O.

In a yet further embodiment, R¹⁰ is—(CH₂)₂—B—O—(CR^(11a)R^(12a))C(O)OR¹³.

In another embodiment of the invention as defined anywhere above, B isphenyl optionally substituted by one or more Z groups.

In a further embodiment B is phenyl optionally substituted by halogen;in a yet further embodiment the halogen is chlorine.

In another embodiment of the invention as defined anywhere above, X isselected from —(SO₂)NR¹⁵—, —NR¹⁵C(O)NR¹⁷—, and O.

In another embodiment of the invention as defined anywhere above, R¹⁵and R¹⁷ are H.

In another embodiment of the invention as defined anywhere above,R^(11a), R^(11b), R^(11c), R^(12a), R^(12b) and R^(12c) are eachindependently selected from H and C₁-C₃ alkyl; or

R^(11a) and R^(12a) together with the carbon atom to which they areattached form a 3-, 4- or 5-membered cycloalkyl group; orR^(11b) and R^(12b) together with the carbon atom to which they areattached form a 3-, 4- or 5-membered cycloalkyl group; orR^(11c) and R^(12c) together with the carbon atom to which they areattached form a 3-, 4- or 5-membered cycloalkyl group.

In another embodiment of the invention as defined anywhere above, m is 0or 1.

In another embodiment of the invention as defined anywhere above, n is 0or 1.

In another embodiment of the invention as defined anywhere above, p is 0or 1.

In another embodiment of the invention as defined anywhere above, thesum of m, n and p is 0, 1, 2, or 3; in a further embodiment the sum is1, 2, or, 3; in a yet further embodiment, the sum is 1 or 2.

In another embodiment of the invention as defined anywhere above whereinX is selected from —(SO₂)NR¹⁵— and —NR¹⁵C(O)NR¹⁷—, the sum of m, n and pis 2.

In another embodiment of the invention as defined anywhere above whereinX is O, and the sum of m, n and p is 1.

In another embodiment of the invention as defined anywhere above, R¹³ isselected from (C₁ alkyl)-C(O)NR²²R²³; (C₁ alkyl)-C(O)OR²³; and (C₂alkyl)-NR²³C(O)R²².

In a further embodiment of the invention as defined anywhere above, R¹³is selected from (C₁ alkyl)-C(O)NR²²R²³ and (C₁ alkyl)-C(O)OR²³.

In a yet further embodiment of the invention as defined anywhere above,R¹³ is (C₁ alkyl)-C(O)NR²²R²³.

In another embodiment of the invention as defined anywhere above,

R²² is selected from H and C₁-C₃ alkyl;R²³ is selected from H; C₁-C₈ alkyl; C₃-C₈ cycloalkyl; C₁-C₄alkoxy-C₁-C₄ alkyl; (C₀-C₄ alkyl)-aryl optionally substituted by one ormore groups selected from C₁-C₆ alkyl, C₁-C₆ alkoxy and halogen; (C₀-C₄alkyl)-3- to 14-membered heterocyclic group, the heterocyclic groupincluding one or more heteroatoms selected from N, O and S, optionallysubstituted by one or more groups selected from halogen, oxo, C₁-C₆alkyl and C(O)C₁-C₆ alkyl; (C₀-C₄ alkyl)-O-aryl optionally substitutedby one or more groups selected from C₁-C₆ alkyl, C₁-C₆ alkoxy andhalogen; and (C₀-C₄ alkyl)-O-3- to 14-membered heterocyclic group, theheterocyclic group including one or more heteroatoms selected from N, Oand S, optionally substituted by one or more groups selected fromhalogen, C₁-C₆ alkyl and C(O)C₁-C₆ alkyl; wherein the alkyl groups areoptionally substituted by one or more halogen atoms, hydroxyl, C₁-C₄alkoxy, C(O)NH₂, C(O)NHC₁-C₆ alkyl or C(O)N(C₁-C₆ alkyl)₂; orR²² and R²³ together with the atom(s) to which they are attached form a5- to 7-membered heterocycloalkyl group, the heterocycloalkyl groupincluding one or more further heteroatoms selected from N, O and S, theheterocycloalkyl group being optionally substituted by one or more Zgroups.

In another embodiment of the invention as defined anywhere above whereinR¹³ is (C₁-C₃ alkyl)-NR²³C(O)R²², R²² andR²³ together with the atoms towhich they are attached form an oxo substituted 5- to 7-memberedheterocycloalkyl group, the heterocycloalkyl group including one or morefurther heteroatoms selected from N, O and S, the heterocycloalkyl groupbeing optionally further substituted by one or more Z groups. In afurther embodiment, R¹³ is (C₁-C₃ alkyl)-NR²³C(O)R²², wherein—NR²³C(O)R²² is represented by formula 3

wherein q is 0, 1 or 2, and the asterisk indicates the point ofattachment to the (C₁-C₃ alkyl)-moiety.

In another embodiment of the invention as defined anywhere above whereinR¹³ is selected from (C₁-C₃ alkyl)-C(O)NR²²R²³ and (C₁-C₃alkyl)-C(O)OR²³;

R²² is selected from H and C₁-C₃ alkyl;R²³ is selected from H; C₁-C₈ alkyl; C₃-C₈ cycloalkyl; C₁-C₄alkoxy-C₁-C₄ alkyl; (C₀-C₄ alkyl)-aryl optionally substituted by one ormore groups selected from C₁-C₆ alkyl, C₁-C₆ alkoxy and halogen; (C₀-C₄alkyl)-3- to 14-membered heterocyclic group, the heterocyclic groupincluding one or more heteroatoms selected from N, O and S, optionallysubstituted by one or more groups selected from halogen, oxo, C₁-C₆alkyl and C(O)C₁-C₆ alkyl; (C₀-C₄ alkyl)-O-aryl optionally substitutedby one or more groups selected from C₁-C₆ alkyl, C₁-C₆ alkoxy andhalogen; and (C₀-C₄ alkyl)-O-3- to 14-membered heterocyclic group, theheterocyclic group including one or more heteroatoms selected from N, Oand S, optionally substituted by one or more groups selected fromhalogen, C₁-C₆ alkyl and C(O)C₁-C₆ alkyl; wherein the alkyl groups areoptionally substituted by one or more halogen atoms, hydroxyl C₁-C₄alkoxy, C(O)NH₂, C(O)NHC₁-C₆ alkyl or C(O)N(C₁-C₆ alkyl)₂; orR²² and R²³ together with the nitrogen to which they are attached form a5- to 7-membered heterocycloalkyl group, the heterocycloalkyl groupincluding one or more further heteroatoms selected from N, O and S, theheterocycloalkyl group being optionally substituted by one or more Zgroups.

In another embodiment of the invention as defined anywhere above, Z ishydroxyl, cyano, nitro, C₁-C₄-alkyl, C₁-C₄-alkoxy, halogen,C₁-C₄-alkylcarbonyl, carboxy, C₁-C₄-alkoxycarbonyl, amino,C₁-C₄-alkylamino, di-C₁-C₄-alkylamino, C₁-C₄-alkylaminocarbonyl,di-C₁-C₄-alkylaminocarbonyl, C₁-C₄-alkylcarbonylamino,C₁-C₄-alkylcarbonyl(C₁-C₄-alkyl)amino, wherein the alkyl and alkoxygroups are each optionally substituted by one or more substituentsselected from hydroxyl, halogen, C₁-C₄-haloalkyl and C₁-C₄-alkoxygroups.

In a yet further embodiment, the invention provides a compound ofFormula Ia

or a pharmaceutically acceptable salt or solvate thereof wherein R¹⁰ isas defined anywhere above in respect of a compound of Formula I.

In a yet further embodiment, the invention provides a compound ofFormula Ib

or a pharmaceutically acceptable salt or solvate thereof wherein B,R^(11a), R^(12a), R^(11b), R^(12b) and R¹³ are as defined anywhere abovein respect of a compound of Formula I.

In a yet further embodiment, the invention provides a compound ofFormula Ic

or a pharmaceutically acceptable salt or solvate thereof wherein B,R^(11a), R^(12a), R^(11b), R^(12b), R¹³ and R¹⁵ and R¹⁷ are as definedanywhere above in respect of a compound of Formula I.

In a yet further embodiment, the invention provides a compound ofFormula Id

or a pharmaceutically acceptable salt or solvate thereof wherein B,R^(11a), R^(12a), and R¹³ are as defined anywhere above in respect of acompound of Formula I.

In another embodiment, individual compounds according to the inventionare those listed in the Examples section below.

In another embodiment of the invention, there is provided a compound ofFormula I which is selected from:

-   3-(3-{2-[(E)-3,5-Diamino-6-chloro-pyrazine-2-carbonylimino]-1,3,8-triaza-spiro[4.5]decane-8-carbonyl}-benzenesulfonylamino)-propionic    acid dipropylcarbamoylmethyl ester;-   [4-(3-{2-[(Z)-3,5-Diamino-6-chloro-pyrazine-2-carbonylimino]-1,3,8-triaza-spiro[4.5]dec-8-yl}-3-oxo-propyl)-phenoxy]-acetic    acid [(2-hydroxy-ethyl)-methyl-carbamoyl]-methyl ester;-   [4-(3-{2-[(E)-3,5-Diamino-6-chloro-pyrazine-2-carbonylimino]-1,3,8-triaza-spiro[4.5]dec-8-yl}-3-oxo-propyl)-phenoxy]-acetic    acid cyclohexyl oxycarbonylmethyl ester;-   3-(3-{2-[(E)-3,5-Diamino-6-chloro-pyrazine-2-carbonylimino]-1,3,8-triaza-spiro[4.5]decane-8-carbonyl}-benzene    sulfonylamino)-propionic acid cyclohexyloxy carbonylmethyl ester;-   [4-(3-{2-[(E)-3,5-Diamino-6-chloro-pyrazine-2-carbonylimino]-1,3,8-triaza-spiro[4.5]dec-8-yl}-3-oxo-propyl)-phenoxy]-acetic    acid dimethylcarbamoylmethyl ester;-   [4-(3-{2-[(E)-3,5-Diamino-6-chloro-pyrazine-2-carbonylimino]-1,3,8-triaza-spiro[4.5]dec-8-yl}-3-oxo-propyl)-phenoxy]-acetic    acid dipropylcarbamoylmethyl ester;-   [4-(3-{2-[(E)-3,5-Diamino-6-chloro-pyrazine-2-carbonylimino]-1,3,8-triaza-spiro[4.5]dec-8-yl}-3-oxo-propyl)-phenoxy]-acetic    acid tert-butoxycarbonylmethyl ester;-   [4-(3-{2-[(E)-3,5-Diamino-6-chloro-pyrazine-2-carbonyl    imino]-1,3,8-triaza-spiro[4.5]dec-8-yl}-3-oxo-propyl)-phenoxy]-acetic    acid benzyloxycarbonylmethyl ester;-   [4-(3-{2-[(E)-3,5-Diamino-6-chloro-pyrazine-2-carbonylimino]-1,3,8-triaza-spiro[4.5]dec-8-yl}-3-oxo-propyl)-phenoxy]-acetic    acid diethylcarbamoylmethyl ester;-   [4-(3-{2-[(E)-3,5-Diamino-6-chloro-pyrazine-2-carbonylimino]-1,3,8-triaza-spiro[4.5]dec-8-yl}-3-oxo-propyl)-phenoxy]-acetic    acid 2-oxo-2-piperidin-1-yl-ethyl ester;-   [2-Chloro-4-(3-{2-[(E)-3,5-diamino-6-chloro-pyrazine-2-carbonylimino]-1,3,8-triaza-spiro[4.5]dec-8-yl}-3-oxo-propyl)-phenoxy]-acetic    acid dipropylcarbamoylmethyl ester;-   3-(3-{2-[(E)-3,5-Diamino-6-chloro-pyrazine-2-carbonylimino]-1,3,8-triaza-spiro[4.5]decane-8-carbonyl}-benzenesulfonylamino)-propionic    acid 2-oxo-2-(2-trifluoromethyl-pyrrolidin-1-yl)-ethyl ester;-   [2-Chloro-4-(3-{2-[(E)-3,5-diamino-6-chloro-pyrazine-2-carbonylimino]-1,3,8-triaza-spiro[4.5]dec-8-yl}-3-oxo-propyl)-phenoxy]-acetic    acid 2-(2-oxo-piperidin-1-yl)-ethyl ester;-   [4-(3-{2-[(E)-3,5-Diamino-6-chloro-pyrazine-2-carbonylimino]-1,3,8-triaza-spiro[4.5]dec-8-yl}-3-oxo-propyl)-phenoxy]-acetic    acid 2-morpholin-4-yl-2-oxo-ethyl ester;-   1-[(3-{2-[(E)-3,5-Diamino-6-chloro-pyrazine-2-carbonylimino]-1,3,8-triaza-spiro[4.5]decane-8-carbonyl}-benzenesulfonylamino)-methyl]-cyclobutanecarboxylic    acid dipropylcarbamoylmethyl ester;-   3-[3-(3-{2-[(E)-3,5-Diamino-6-chloro-pyrazine-2-carbonylimino]-1,3,8-triaza-spiro[4.5]decane-8-carbonyl}-phenyl)-ureido]-propionic    acid dipropylcarbamoylmethyl ester; and-   1-[(3-{2-[(E)-3,5-Diamino-6-chloro-pyrazine-2-carbonylimino]-1,3,8-triaza-spiro[4.5]decane-8-carbonyl}-benzenesulfonylamino)-methyl]-cyclobutanecarboxylic    acid-   2-oxo-2-(2-trifluoromethyl-pyrrolidin-1-yl)-ethyl ester;    or a pharmaceutically acceptable salt or solvate thereof.

In the embodiments mentioned herein, where only certain variables aredefined, it is intended that the remainder of the variables are asdefined in any embodiment herein. Thus, the invention provides for thecombination of limited or optional definitions of variables.

The following terms as used herein are intended to have the followingmeanings:

“Optionally substituted” as used herein means the group referred to canbe unsubstituted, or substituted at one or two or three positions by anyone or any combination of the radicals listed thereafter.

“Halo” or “halogen” as used herein means fluorine, chlorine, bromine oriodine.

“C₁-C₃ alkyl”, “C₁-C₆ alkyl”, “C₁-C₈ alkyl” and the like, as usedherein, denotes a straight chain or branched alkyl group that containsone to three, six or eight (or the relevant number) carbon atoms andwhich may be substituted as defined.

“Aryl”, as used herein, represents an aromatic carbocyclic ring systemhaving 6 to 15 carbon atoms. It can be monocyclic, bicyclic ortricyclic, and may be optionally substituted as defined. Examples ofC₆-C₁₅-aryl groups include but are not limited to phenyl, phenylene,benzenetriyl, indanyl, naphthyl, naphthylene, naphthalenetriyl andanthracenyl.

“Heterocyclyl” or “heterocyclic” refers to a 4- to 14-memberedheterocyclic ring system containing at least one ring heteroatomselected from the group consisting of nitrogen, oxygen and sulphur,which may be saturated, partially saturated or aromatic (i.e.heteroaryl). Examples of 4- to 14-membered heterocyclic groups includebut are not limited to furan, azetidine, pyrrole, pyrrolidine, pyrazole,imidazole, triazole, isotriazole, tetrazole, thiadiazole, isothiazole,oxadiazole, pyridine, piperidine, pyrazine, oxazole, isoxazole,pyrazine, pyridazine, pyrimidine, piperazine, pyrrolidine,pyrrolidinone, pyridinone, morpholine, triazine, oxazine,tetrahydrofuran, tetrahydrothiophene, tetrahydrothiopyran,tetrahydropyran, 1,4-dioxane, 1,4-oxathiane, indazole, quinoline,quinazoline, quinoxaline, indole, indoline, thiazole, thiophene,isoquinoline, isoindole, isoindoline, benzothiophene, benzoxazole,benzisoxazole, benzothiazole, benzisothiazole, benzofuran,dihydrobenzofuran, dihydroisobenzofuran, benzodioxole, benzimidazole,benzotriazole, pyrazolopyridine, pyrazolopyrimidine, imidazopyridine,purine, naphthyridine or tetrahydronaphthyridine. “Heterocyclyl” or“heterocyclic” also includes bridged heterocyclic groups such as3-hydroxy-8-aza-bicyclo[3.2.1]oct-8-yl and fused ring systems such aspyridopyrimidine. The 4- to 14-membered heterocyclic group can beunsubstituted or substituted.

“Heterocyclyl” includes heteroaryl and heterocycloalkyl groups.

“Heteroaryl” is an aromatic ring system containing from 5 to 15 ringatoms one or more of which are heteroatoms selected from O, N or S.Preferably there are one or two heteroatoms. Heteroaryl (heterocyclicaryl) represents, for example: pyridyl, indolyl, isoindolyl, indazolyl,quinoxalinyl, quinazolinyl, quinolinyl, isoquinolinyl, naphthryridinyl,pyridopyrimidinyl, benzothienyl, benzofuranyl, benzopyranyl,benzothiopyranyl, benzotriazolyl, pyrazolopyridinyl, furanyl, pyrrolyl,thiazolyl, oxazolyl, isoxazolyl, triazolyl, tetrazolyl, pyrazolyl,imidazolyl, thienyl. The heteroaryl group can be substituted orunsubstituted.

“C₃-C₁₀-cycloalkyl” denotes a fully saturated carbocyclic ring having 3to 10 ring carbon atoms, for example a monocyclic group such as acyclopropyl, cyclobutyl, cyclopentyl or cyclohexyl, cycloheptyl,cyclooctyl, cyclononyl or cyclodecyl, or a bicyclic group such asbicycloheptyl or bicyclooctyl. Different numbers of carbon atoms may bespecified, with the definition being amended accordingly. The cycloalkylgroup can be substituted or unsubstituted.

“C₅-C₁₀-cycloalkenyl” denotes a partially saturated carbocyclic ringhaving 5 to 10 ring carbon atoms, for example a monocyclic group such asa cyclopentenyl or cyclohexenyl, cycloheptenyl, cyclooctenyl orcyclononenyl, or a bicyclic group such as bicycloheptenyl orbicyclooctenyl. The ring or ring system may contain more than onecarbon-carbon double bond. Different numbers of carbon atoms may bespecified, with the definition being amended accordingly. Thecycloalkenyl group can be substituted or unsubstituted.

“C₁-C₈-haloalkyl” as used herein denotes C₁-C₈-alkyl as hereinbeforedefined substituted by one or more halogen atoms, preferably one, two orthree halogen atoms. Different numbers of carbon atoms may be specified,with the definition being amended accordingly.

“C₁-C₈-alkylamino” as used herein denote amino substituted by one or twoC₁-C₈-alkyl groups as hereinbefore defined, which may be the same ordifferent. Different numbers of carbon atoms may be specified, with thedefinition being amended accordingly.

“C₁-C₈-alkoxy” as used herein denotes straight chain or branched alkoxythat contains 1 to 8 carbon atoms. Different numbers of carbon atoms maybe specified, with the definition being amended accordingly.

Throughout this specification and in the claims that follow, unless thecontext requires otherwise, the word “comprise”, or variations such as“comprises” or “comprising”, should be understood to imply the inclusionof a stated integer or step or group of integers or steps but not theexclusion of any other integer or step or group of integers or steps.

As used herein, the term “pharmaceutically acceptable salts” refers tosalts that retain the biological effectiveness and properties of thecompounds of this invention and, which typically are not biologically orotherwise undesirable. In many cases, the compounds of the presentinvention are capable of forming acid and/or base salts by virtue of thepresence of amino and/or carboxyl groups or groups similar thereto.

Pharmaceutically acceptable acid addition salts can be formed withinorganic acids and organic acids, e.g., acetate, aspartate, benzoate,besylate, bromide/hydrobromide, bicarbonate/carbonate,bisulfate/sulfate, camphorsulfornate, chloride/hydrochloride,chlortheophyllonate, citrate, ethandisulfonate, fumarate, gluceptate,gluconate, glucuronate, hippurate, hydroiodide/iodide, isethionate,lactate, lactobionate, laurylsulfate, malate, maleate, malonate,mandelate, mesylate, methylsulphate, naphthoate, napsylate, nicotinate,nitrate, octadecanoate, oleate, oxalate, palmitate, pamoate,phosphate/hydrogen phosphate/dihydrogen phosphate, polygalacturonate,propionate, stearate, succinate, sulfosalicylate, tartrate, tosylate andtrifluoroacetate salts.

Inorganic acids from which salts can be derived include, for example,hydrochloric acid, hydrobromic acid, sulfuric acid, nitric acid,phosphoric acid, and the like.

Organic acids from which salts can be derived include, for example,acetic acid, propionic acid, glycolic acid, oxalic acid, maleic acid,malonic acid, succinic acid, fumaric acid, tartaric acid, citric acid,benzoic acid, mandelic acid, methanesulfonic acid, ethanesulfonic acid,toluenesulfonic acid, sulfosalicylic acid, and the like.Pharmaceutically acceptable base addition salts can be formed withinorganic and organic bases.

Inorganic bases from which salts can be derived include, for example,ammonium salts and metals from columns I to XII of the periodic table.In certain embodiments, the salts are derived from sodium, potassium,ammonium, calcium, magnesium, iron, silver, zinc, and copper;particularly suitable salts include ammonium, potassium, sodium, calciumand magnesium salts.

Organic bases from which salts can be derived include, for example,primary, secondary, and tertiary amines, substituted amines includingnaturally occurring substituted amines, cyclic amines, basic ionexchange resins, and the like. Certain organic amines includeisopropylamine, benzathine, cholinate, diethanolamine, diethylamine,lysine, meglumine, piperazine and tromethamine.

The pharmaceutically acceptable salts of the present invention can besynthesized from a parent compound, a basic or acidic moiety, byconventional chemical methods. Generally, such salts can be prepared byreacting free acid forms of these compounds with a stoichiometric amountof the appropriate base (such as Na, Ca, Mg, or K hydroxide, carbonate,bicarbonate or the like), or by reacting free base forms of thesecompounds with a stoichiometric amount of the appropriate acid. Suchreactions are typically carried out in water or in an organic solvent,or in a mixture of the two. Generally, use of non-aqueous media likeether, ethyl acetate, ethanol, isopropanol, or acetonitrile isdesirable, where practicable. Lists of additional suitable salts can befound, e.g., in “Remington's Pharmaceutical Sciences”, 20th ed., MackPublishing Company, Easton, Pa., (1985); and in “Handbook ofPharmaceutical Salts: Properties, Selection, and Use” by Stahl andWermuth (Wiley-VCH, Weinheim, Germany, 2011).

Furthermore, the compounds of the present invention, including theirsalts, can also be obtained in the form of their hydrates, or includeother solvents used for their crystallization.

Compounds of the invention, i.e. compounds of formula (I) that containgroups capable of acting as donors and/or acceptors for hydrogen bondsmay be capable of forming co-crystals with suitable co-crystal formers.These co-crystals may be prepared from compounds of formula (I) by knownco-crystal forming procedures. Such procedures include grinding,heating, co-subliming, co-melting, or contacting in solution compoundsof formula (I) with the co-crystal former under crystallizationconditions and isolating co-crystals thereby formed. Suitable co-crystalformers include those described in WO 2004/078163. Hence the inventionfurther provides co-crystals comprising a compound of formula (I).

As used herein, the term “an optical isomer” or “a stereoisomer” refersto any of the various stereo isomeric configurations which may exist fora given compound of the present invention and includes geometricisomers. It is understood that a substituent may be attached at a chiralcenter of a carbon atom. Therefore, the invention includes enantiomers,diastereomers or racemates of the compound. “Enantiomers” are a pair ofstereoisomers that are non-superimposable mirror images of each other. A1:1 mixture of a pair of enantiomers is a “racemic” mixture. The term isused to designate a racemic mixture where appropriate.“Diastereoisomers” are stereoisomers that have at least two asymmetricatoms, but which are not mirror-images of each other. The absolutestereochemistry is specified according to the Cahn-Ingold-Prelog R-Ssystem. When a compound is a pure enantiomer the stereochemistry at eachchiral carbon may be specified by either R or S. Resolved compoundswhose absolute configuration is unknown can be designated (+) or (−)depending on the direction (dextro- or levorotatory) which they rotateplane polarized light at the wavelength of the sodium D line. Certain ofthe compounds described herein contain one or more asymmetric centers oraxes and may thus give rise to enantiomers, diastereomers, and otherstereoisomeric forms that may be defined, in terms of absolutestereochemistry, as (R)- or (S)-. The present invention is meant toinclude all such possible isomers, including racemic mixtures, opticallypure forms and intermediate mixtures. Optically active (R)- and(S)-isomers may be prepared using chiral synthons or chiral reagents, orresolved using conventional techniques. If the compound contains adouble bond, the substituent may be E or Z configuration. If thecompound contains a disubstituted cycloalkyl, the cycloalkyl substituentmay have a cis- or trans-configuration. All tautomeric forms are alsointended to be included. Tautomers are one of two or more structuralisomers that exist in equilibrium and are readily converted from oneisomeric form to another. More specifically, for example, compounds ofFormula Ia may exist in one or both of the following tautomeric forms:

Examples of tautomers include but are not limited to those compoundsdefined in the claims.

Any asymmetric atom (e.g., carbon or the like) of the compound(s) of thepresent invention can be present in racemic or enantiomericallyenriched, for example the (R)-, (S)- or (R,S)-configuration. In certainembodiments, each asymmetric atom has at least 50% enantiomeric excess,at least 60% enantiomeric excess, at least 70% enantiomeric excess, atleast 80% enantiomeric excess, at least 90% enantiomeric excess, atleast 95% enantiomeric excess, or at least 99% enantiomeric excess inthe (R)- or (S)-configuration. Substituents at atoms with unsaturatedbonds may, if possible, be present in cis-(Z)- or trans-(E)-form.

Accordingly, as used herein a compound of the present invention can bein the form of one of the possible isomers, rotamers, atropisomers,tautomers or mixtures thereof, for example, as substantially puregeometric (cis or trans) isomers, diastereomers, optical isomers(antipodes), racemates or mixtures thereof.

Any resulting mixtures of isomers can be separated on the basis of thephysicochemical differences of the constituents, into the pure orsubstantially pure geometric or optical isomers, diastereomers,racemates, for example, by chromatography and/or fractionalcrystallization.

Any resulting racemates of final products or intermediates can beresolved into the optical antipodes by known methods, e.g., byseparation of the diastereomeric salts thereof, obtained with anoptically active acid or base, and liberating the optically activeacidic or basic compound. In particular, a basic moiety may thus beemployed to resolve the compounds of the present invention into theiroptical antipodes, e.g., by fractional crystallization of a salt formedwith an optically active acid, e.g., tartaric acid, dibenzoyl tartaricacid, diacetyl tartaric acid, di-O,O′-p-toluoyl tartaric acid, mandelicacid, malic acid or camphor-10-sulfonic acid. Racemic products can alsobe resolved by chiral chromatography, e.g., high pressure liquidchromatography (HPLC) using a chiral adsorbent.

Since the compounds of the invention are intended for use inpharmaceutical compositions it will readily be understood that they areeach preferably provided in substantially pure form, for example atleast 60% pure, more suitably at least 75% pure and preferably at least85%, especially at least 98% pure (% are on a weight for weight basis).Impure preparations of the compounds may be used for preparing the morepure forms used in the pharmaceutical compositions; these less purepreparations of the compounds should contain at least 1%, more suitablyat least 5% and preferably from 10 to 59% of a compound of theinvention.

Compounds of the present invention are either obtained in the free form,as a salt thereof, or as prodrug derivatives thereof.

When both a basic group and an acid group are present in the samemolecule, the compounds of the present invention may also form internalsalts, e.g., zwitterionic molecules.

The present invention also provides pro-drugs of the compounds of thepresent invention that converts in vivo to the compounds of the presentinvention. A pro-drug is an active or inactive compound that is modifiedchemically through in vivo physiological action, such as hydrolysis,metabolism and the like, into a compound of this invention followingadministration of the prodrug to a subject. The suitability andtechniques involved in making and using pro-drugs are well known bythose skilled in the art. Prodrugs can be conceptually divided into twonon-exclusive categories, bioprecursor prodrugs and carrier prodrugs.See The Practice of Medicinal Chemistry, Ch. 31-32 (Ed. Wermuth,Academic Press, San Diego, Calif., 2001). Generally, bioprecursorprodrugs are compounds, which are inactive or have low activity comparedto the corresponding active drug compound, that contain one or moreprotective groups and are converted to an active form by metabolism orsolvolysis. Both the active drug form and any released metabolicproducts should have acceptably low toxicity.

Carrier prodrugs are drug compounds that contain a transport moiety,e.g., that improve uptake and/or localized delivery to a site(s) ofaction. Desirably for such a carrier prodrug, the linkage between thedrug moiety and the transport moiety is a covalent bond, the prodrug isinactive or less active than the drug compound, and any releasedtransport moiety is acceptably non-toxic. For prodrugs where thetransport moiety is intended to enhance uptake, typically the release ofthe transport moiety should be rapid. In other cases, it is desirable toutilize a moiety that provides slow release, e.g., certain polymers orother moieties, such as cyclodextrins. Carrier prodrugs can, forexample, be used to improve one or more of the following properties:increased lipophilicity, increased duration of pharmacological effects,increased site-specificity, decreased toxicity and adverse reactions,and/or improvement in drug formulation (e.g., stability, watersolubility, suppression of an undesirable organoleptic or physiochemicalproperty). For example, lipophilicity can be increased by esterificationof (a) hydroxyl groups with lipophilic carboxylic acids (e.g., acarboxylic acid having at least one lipophilic moiety), or (b)carboxylic acid groups with lipophilic alcohols (e.g., an alcohol havingat least one lipophilic moiety, for example aliphatic alcohols).

Exemplary prodrugs are, e.g., esters of free carboxylic acids and S-acylderivatives of thiols and O-acyl derivatives of alcohols or phenols,wherein acyl has a meaning as defined herein. Suitable prodrugs areoften pharmaceutically acceptable ester derivatives convertible bysolvolysis under physiological conditions to the parent carboxylic acid,e.g., lower alkyl esters, cycloalkyl esters, lower alkenyl esters,benzyl esters, mono- or di-substituted lower alkyl esters, such as theω-(amino, mono- or di-lower alkylamino, carboxy, loweralkoxycarbonyl)-lower alkyl esters, the α-(lower alkanoyloxy, loweralkoxycarbonyl or di-lower alkylaminocarbonyl)-lower alkyl esters, suchas the pivaloyloxymethyl ester and the like conventionally used in theart. In addition, amines have been masked as arylcarbonyloxymethylsubstituted derivatives which are cleaved by esterases in vivo releasingthe free drug and formaldehyde (Bundgaard, J. Med. Chem. 2503 (1989)).Moreover, drugs containing an acidic NH group, such as imidazole, imide,indole and the like, have been masked with N-acyloxymethyl groups(Bundgaard, Design of Prodrugs, Elsevier (1985)). Hydroxy groups havebeen masked as esters and ethers. EP 039,051 (Sloan and Little)discloses Mannich-base hydroxamic acid prodrugs, their preparation anduse.

Any formula given herein is also intended to represent unlabeled formsas well as isotopically labeled forms of the compounds. Isotopicallylabeled compounds have structures depicted by the formulas given hereinexcept that one or more atoms are replaced by an atom having a selectedatomic mass or mass number. Examples of isotopes that can beincorporated into compounds of the invention include isotopes ofhydrogen, carbon, nitrogen, oxygen, phosphorous, fluorine, and chlorine,such as ²H, ³H, ¹¹C, ¹³C, ¹⁴C, ¹⁵N, ¹⁸F ³¹P, ³²P, ³⁵S, ³⁶Cl, ¹²⁵Irespectively. The invention includes various isotopically labeledcompounds as defined herein, for example those into which radioactiveisotopes, such as ³H, ¹³C, and ¹⁴C, are present. Such isotopicallylabelled compounds are useful in metabolic studies (with ¹⁴C), reactionkinetic studies (with, for example ²H or ³H), detection or imagingtechniques, such as positron emission tomography (PET) or single-photonemission computed tomography (SPECT) including drug or substrate tissuedistribution assays, or in radioactive treatment of patients. Inparticular, an ¹⁸F or labeled compound may be particularly desirable forPET or SPECT studies. Isotopically labeled compounds of this inventionand prodrugs thereof can generally be prepared by carrying out theprocedures disclosed in the schemes or in the examples and preparationsdescribed below by substituting a readily available isotopically labeledreagent for a non-isotopically labeled reagent.

Further, substitution with heavier isotopes, particularly deuterium(i.e., ²H or D) may afford certain therapeutic advantages resulting fromgreater metabolic stability, for example increased in vivo half-life orreduced dosage requirements or an improvement in therapeutic index. Itis understood that deuterium in this context is regarded as asubstituent of a compound of the formula (I). The concentration of sucha heavier isotope, specifically deuterium, may be defined by theisotopic enrichment factor. The term “isotopic enrichment factor” asused herein means the ratio between the isotopic abundance and thenatural abundance of a specified isotope. If a substituent in a compoundof this invention is denoted deuterium, such compound has an isotopicenrichment factor for each designated deuterium atom of at least 3500(52.5% deuterium incorporation at each designated deuterium atom), atleast 4000 (60% deuterium incorporation), at least 4500 (67.5% deuteriumincorporation), at least 5000 (75% deuterium incorporation), at least5500 (82.5% deuterium incorporation), at least 6000 (90% deuteriumincorporation), at least 6333.3 (95% deuterium incorporation), at least6466.7 (97% deuterium incorporation), at least 6600 (99% deuteriumincorporation), or at least 6633.3 (99.5% deuterium incorporation).

Isotopically-labeled compounds of formula (I) can generally be preparedby conventional techniques known to those skilled in the art or byprocesses analogous to those described in the accompanying Examples andPreparations using an appropriate isotopically-labeled reagents in placeof the non-labeled reagent previously employed. Pharmaceuticallyacceptable solvates in accordance with the invention include thosewherein the solvent of crystallization may be isotopically substituted,e.g. D₂O, d₆-acetone, d₆-DMSO.

Synthesis

The compounds of the invention may be synthesized by the generalsynthetic routes below, specific examples of which are described in moredetail in the Examples.

Compounds of formula I may be prepared according to Scheme 1.

Alternatively, compounds of formula I may be prepared according toScheme 2.

wherein R^(10a) is—(C₀-C₃alkylene)-B—X—(CR^(11a)R^(12a))_(m)—(CR^(11b)R^(12b))_(n)—(CR^(11c)R^(12c))_(p)—C(O)OH,and is L is a suitable leaving group.

The above general schemes may be used to prepare compounds of Formula I.The desired specific compounds can be prepared by selecting theappropriate starting materials, reactants and reaction conditions.

The starting materials and reagents in the above scheme are all eitheravailable commercially or can be prepared following literatureprecedents. In particular compounds of Formula II and compounds ofFormula IV may be prepared as described in international patentapplication published as WO2009074575, which document is incorporatedherein by reference.

Within the scope of this text, only a readily removable group that isnot a constituent of the particular desired end product of the compoundsof the present invention is designated a “protecting group”, unless thecontext indicates otherwise. The protection of functional groups by suchprotecting groups, the protecting groups themselves, and their cleavagereactions are described for example in standard reference works, such asJ. F. W. McOmie, “Protective Groups in Organic Chemistry”, Plenum Press,London and New York 1973, in T. W. Greene and P. G. M. Wuts, “ProtectiveGroups in Organic Synthesis”, Third edition, Wiley, New York 1999, in“The Peptides”; Volume 3 (editors: E. Gross and J. Meienhofer), AcademicPress, London and New York 1981, in “Methoden der organischen Chemie”(Methods of Organic Chemistry), Houben Weyl, 4th edition, Volume 15/I,Georg Thieme Verlag, Stuttgart 1974, in H.-D. Jakubke and H. Jeschkeit,“Aminosauren, Peptide, Proteine” (Amino acids, Peptides, Proteins),Verlag Chemie, Weinheim, Deerfield Beach, and Basel 1982, and in JochenLehmann, “Chemie der Kohlenhydrate: Monosaccharide and Derivate”(Chemistry of Carbohydrates: Monosaccharides and Derivatives), GeorgThieme Verlag, Stuttgart 1974. A characteristic of protecting groups isthat they can be removed readily (i.e. without the occurrence ofundesired secondary reactions) for example by solvolysis, reduction,photolysis or alternatively under physiological conditions (e.g. byenzymatic cleavage). Salts of compounds of the present invention havingat least one salt-forming group may be prepared in a manner known tothose skilled in the art. For example, salts of compounds of the presentinvention having acid groups may be formed, for example, by treating thecompounds with metal compounds, such as alkali metal salts of suitableorganic carboxylic acids, e.g. the sodium salt of 2-ethylhexanoic acid,with organic alkali metal or alkaline earth metal compounds, such as thecorresponding hydroxides, carbonates or hydrogen carbonates, such assodium or potassium hydroxide, carbonate or hydrogen carbonate, withcorresponding calcium compounds or with ammonia or a suitable organicamine, stoichiometric amounts or only a small excess of the salt-formingagent preferably being used. Acid addition salts of compounds of thepresent invention are obtained in customary manner, e.g. by treating thecompounds with an acid or a suitable anion exchange reagent. Internalsalts of compounds of the present invention containing acid and basicsalt-forming groups, e.g. a free carboxy group and a free amino group,may be formed, e.g. by the neutralisation of salts, such as acidaddition salts, to the isoelectric point, e.g. with weak bases, or bytreatment with ion exchangers. Salts can be converted into the freecompounds in accordance with methods known to those skilled in the art.Metal and ammonium salts can be converted, for example, by treatmentwith suitable acids, and acid addition salts, for example, by treatmentwith a suitable basic agent.

Mixtures of isomers obtainable according to the invention can beseparated in a manner known to those skilled in the art into theindividual isomers; diastereoisomers can be separated, for example, bypartitioning between polyphasic solvent mixtures, recrystallisationand/or chromatographic separation, for example over silica gel or bye.g. medium pressure liquid chromatography over a reversed phase column,and racemates can be separated, for example, by the formation of saltswith optically pure salt-forming reagents and separation of the mixtureof diastereoisomers so obtainable, for example by means of fractionalcrystallisation, or by chromatography over optically active columnmaterials.

Intermediates and final products can be worked up and/or purifiedaccording to standard methods, e.g. using chromatographic methods,distribution methods, (re-) crystallization, and the like.

The following applies in general to all processes mentioned hereinbefore and hereinafter.

All the above-mentioned process steps can be carried out under reactionconditions that are known to those skilled in the art, including thosementioned specifically, in the absence or, customarily, in the presenceof solvents or diluents, including, for example, solvents or diluentsthat are inert towards the reagents used and dissolve them, in theabsence or presence of catalysts, condensation or neutralizing agents,for example ion exchangers, such as cation exchangers, e.g. in the H+form, depending on the nature of the reaction and/or of the reactants atreduced, normal or elevated temperature, for example in a temperaturerange of from about −100° C. to about 190° C., including, for example,from approximately −80° C. to approximately 150° C., for example at from−80 to −60° C., at room temperature, at from −20 to 40° C. or at refluxtemperature, under atmospheric pressure or in a closed vessel, whereappropriate under pressure, and/or in an inert atmosphere, for exampleunder an argon or nitrogen atmosphere.

At all stages of the reactions, mixtures of isomers that are formed canbe separated into the individual isomers, for example diastereoisomersor enantiomers, or into any desired mixtures of isomers, for exampleracemates or mixtures of diastereoisomers, for example analogously tothe methods described under “Additional process steps”.

The solvents from which those solvents that are suitable for anyparticular reaction may be selected include those mentioned specificallyor, for example, water, esters, such as lower alkyl-lower alkanoates,for example ethyl acetate, ethers, such as aliphatic ethers, for examplediethyl ether, or cyclic ethers, for example tetrahydrofuran or dioxane,liquid aromatic hydrocarbons, such as benzene or toluene, alcohols, suchas methanol, ethanol or 1- or 2-propanol, nitriles, such asacetonitrile, halogenated hydrocarbons, such as methylene chloride orchloroform, acid amides, such as dimethylformamide or dimethylacetamide, bases, such as heterocyclic nitrogen bases, for examplepyridine or N-methylpyrrolidin-2-one, carboxylic acid anhydrides, suchas lower alkanoic acid anhydrides, for example acetic anhydride, cyclic,linear or branched hydrocarbons, such as cyclohexane, hexane orisopentane, methycyclohexane, or mixtures of those solvents, for exampleaqueous solutions, unless otherwise indicated in the description of theprocesses. Such solvent mixtures may also be used in working up, forexample by chromatography or partitioning.

The compounds, including their salts, may also be obtained in the formof hydrates, or their crystals may, for example, include the solventused for crystallization. Different crystalline forms may be present.

The invention relates also to those forms of the process in which acompound obtainable as an intermediate at any stage of the process isused as starting material and the remaining process steps are carriedout, or in which a starting material is formed under the reactionconditions or is used in the form of a derivative, for example in aprotected form or in the form of a salt, or a compound obtainable by theprocess according to the invention is produced under the processconditions and processed further in situ. All starting materials,building blocks, reagents, acids, bases, dehydrating agents, solventsand catalysts utilized to synthesize the compounds of the presentinvention are either commercially available or can be produced byorganic synthesis methods known to one of ordinary skill in the art(Houben-Weyl 4^(th) Ed. 1952, Methods of Organic Synthesis, Thieme,Volume 21).

As a further aspect of the present invention, there is also provided aprocess for the preparation of compounds of formula I or apharmaceutically acceptable salt or solvate thereof.

According to a further aspect of the invention there is provided aprocess of preparing a compound of formula I or a pharmaceuticallyacceptable salt or solvate thereof comprising the step of:

-   -   (a) reacting a compound of formula II

wherein R¹, R², R³, R⁴, R⁵, R⁶, R⁷, R⁸ and R⁹ are as defined anywhereabove, with a compound of formula III

wherein R¹⁰ are as defined anywhere above, under convention reactionconditions for acid-amine coupling; or

-   -   (b) reacting a compound of formula IV

with a compound R¹³L of formula V under convention reaction conditionswherein R¹, R², R³, R⁴, R⁵, R⁶, R⁷, R⁸, R⁹ and R¹³ are as definedanywhere above; R^(10a) is—(C₀-C₃alkylene)-B—X—(CR^(11a)R^(12a))_(m)—(CR^(11b)R^(12b))_(n)—(CR^(11c)R^(12c))_(p)—C(O)OHwherein B, X, R^(11a), R^(11b), R^(11c), R^(12a), R^(112b), R^(12c), m,n, and p are as defined anywhere above; and L is a leaving group.

The invention further includes any variant of the present processes, inwhich an intermediate product obtainable at any stage thereof is used asstarting material and the remaining steps are carried out, or in whichthe starting materials are formed in situ under the reaction conditions,or in which the reaction components are used in the form of their saltsor optically pure antipodes.

Compounds of the invention and intermediates can also be converted intoeach other according to methods generally known to those skilled in theart.

The agents of the invention act to blockade the epithelial sodiumchannel (ENaC). Having regard to their blockade of the epithelial sodiumchannel (ENaC), compounds of formula (I), in free or pharmaceuticallyacceptable salt form, hereinafter alternately referred to as “agents ofthe invention”, are useful in the treatment of conditions which respondto the blockade of the epithelial sodium channel, particularlyconditions benefiting from mucosal hydration.

Diseases mediated by blockade of the epithelial sodium channel, includediseases associated with the regulation of fluid volumes acrossepithelial membranes. For example, the volume of airway surface liquidis a key regulator of mucociliary clearance and the maintenance of lunghealth. The blockade of the epithelial sodium channel will promote fluidaccumulation on the mucosal side of the airway epithelium therebypromoting mucus clearance and preventing the accumulation of mucus andsputum in respiratory tissues (including lung airways). Such diseasesinclude respiratory diseases, such as cystic fibrosis, primary ciliarydyskinesia, chronic bronchitis, chronic obstructive pulmonary disease(COPD), asthma, respiratory tract infections (acute and chronic; viraland bacterial) and lung carcinoma. Diseases mediated by blockade of theepithelial sodium channel also include diseases other than respiratorydiseases that are associated with abnormal fluid regulation across anepithelium, perhaps involving abnormal physiology of the protectivesurface liquids on their surface, e.g., xerostomia (dry mouth) orkeratoconjunctivitis sire (dry eye). Furthermore, blockade of theepithelial sodium channel in the kidney could be used to promotediuresis and thereby induce a hypotensive effect.

Treatment in accordance with the invention may be symptomatic orprophylactic, in particular symptomatic.

Thus in a further aspect the invention includes an agent of theinvention for use as a pharmaceutical.

Therefore according to a further aspect, the invention provides an agentof the invention for treating or preventing a disease or conditionmediated by blockade of the epithelial sodium channel.

Therefore according to a further aspect, the invention provides the useof an agent of the invention in the manufacture of a medicament for thetreatment or prevention of a disease or condition mediated by blockadeof the epithelial sodium channel.

Therefore according to a further aspect, the invention provides a methodfor preventing or treating a disease or condition mediated by blockadeof the epithelial sodium channel in which an effective amount of anagent of the invention is administered to a patient in need of suchtreatment.

Asthma includes both intrinsic (non-allergic) asthma and extrinsic(allergic) asthma, mild asthma, moderate asthma, severe asthma,bronchitic asthma, exercise-induced asthma, occupational asthma andasthma induced following bacterial infection. Treatment of asthma isalso to be understood as embracing treatment of subjects, e.g., of lessthan 4 or 5 years of age, exhibiting wheezing symptoms and diagnosed ordiagnosable as “wheezy infants”, an established patient category ofmajor medical concern and now often identified as incipient orearly-phase asthmatics. (For convenience this particular asthmaticcondition is referred to as “wheezy-infant syndrome”)

Prophylactic efficacy in the treatment of asthma will be evidenced byreduced frequency or severity of symptomatic attack, e.g., of acuteasthmatic or bronchoconstrictor attack, improvement in lung function orimproved airways hyperreactivity. It may further be evidenced by reducedrequirement for other, symptomatic therapy, i.e., therapy for orintended to restrict or abort symptomatic attack when it occurs, e.g.,anti-inflammatory (e.g., cortico-steroid) or bronchodilatory.Prophylactic benefit in asthma may, in particular, be apparent insubjects prone to “morning dipping”. “Morning dipping” is a recognizedasthmatic syndrome, common to a substantial percentage of asthmatics andcharacterized by asthma attack, e.g., between the hours of about 4-6 am,i.e., at a time normally substantially distant from any previouslyadministered symptomatic asthma therapy.

Chronic obstructive pulmonary disease includes chronic bronchitis ordyspnea associated therewith, emphysema, as well as exacerbation ofairways hyperreactivity consequent to other drug therapy, in particular,other inhaled drug therapy. The invention is also applicable to thetreatment of bronchitis of whatever type or genesis including, e.g.,acute, arachidic, catarrhal, croupus, chronic or phthinoid bronchitis.The agents of the invention may also be useful as acid-sensing ionchannel (ASIC) blockers. Thus they may be useful in the treatment ofconditions which respond to the blockade of the acid-sensing ionchannel.

The suitability of epithelial sodium channel blocker as a treatment of adisease benefiting from mucosal hydration, may be tested by determiningthe inhibitory effect of the channel blocker on ENaC in a suitablecell-based assay. For example single cells or confluent epithelia,endogenously expressing or engineered to overexpress ENaC can be used toassess channel function using electrophysiological techniques or ionflux studies. See methods described in: Hirsh et al., J Pharm Exp Ther(2004); Moody et al., Am J Physiol Cell Physiol (2005).

In accordance with the foregoing, the invention also provides as afurther aspect a method for preventing or treating a conditionresponsive to blockade of the epithelial sodium channel, e.g., diseasesassociated with the regulation of fluid volumes across epithelialmembranes, particularly an obstructive airways disease, which comprisesadministering to a subject, particularly a human subject, in needthereof a compound of formula (I), or a pharmaceutically acceptable saltor solvate thereof.

In another aspect the invention provides a compound of formula (I), infree form or or a pharmaceutically acceptable salt or solvate thereof,for preventing or treating a condition responsive to blockade of theepithelial sodium channel, particularly an obstructive airways disease,e.g., cystic fibrosis and COPD.

In another aspect the invention provides the use of a compound offormula (I), in free form or a pharmaceutically acceptable salt orsolvate thereof, in the manufacture of a medicament for the preventionor treatment of a condition responsive to blockade of the epithelialsodium channel, particularly an obstructive airways disease, e.g.,cystic fibrosis and COPD

The compounds of the invention have good ENaC blocker activity and maybe tested in the following assays.

Cell Culture

Human Bronchial Epithelial cells (HBECs) (Cambrex) were cultured underair-liquid interface conditions to provide a well differentiatedmucociliary phenotype.

HBECs were cultured using a modification of the method described by Grayand colleagues (Gray et al., 1996). Cells were seeded in plastic T-162flasks and were grown in bronchial epithelial cell growth medium (BEGM;Cambrex) supplemented with bovine pituitary extract (52 μg/mL),hydrocortisone (0.5 μg/mL), human recombinant epidermal growth factor(0.5 ng/mL), epinephrine (0.5 μg/mL), transferrin (10 μg/mL), insulin (5μg/mL), retinoic acid (0.1 μg/mL), triiodothyronine (6.5 μg/mL),gentamycin (50 μg/mL) and amphotericin B (50 ng/mL). Medium was changedevery 48 hours until cells were 90% confluent. Cells were then passagedand seeded (8.25×10⁵ cells/insert) on polycarbonate Snapwell inserts(Costar) in differentiation media containing 50% DMEM in BEGM with thesame supplements as above but without triiodothyronine and a finalretinoic acid concentration of 50 nM (all-trans retinoic acid). Cellswere maintained submerged for the first 7 days in culture, after whichtime they were exposed to an apical air interface for the remainder ofthe culture period. At this time, media was changed to DMEM:F12 mediacontaining 2% v/v Ultroser G for the remainder of culture. AmphotericinB was removed from all media 3 feeds prior to use in the UssingChambers. Cells were used between days 7 and 21 after establishment ofthe apical-air interface. At all stages of culture, cells weremaintained at 37° C. in 5% CO₂ in an air incubator.

Short Circuit Current (ISC) Measurements

Snapwell inserts were mounted in Vertical Diffusion Chambers (Costar)and were bathed with continuously gassed Ringer solution (5% CO₂ in O₂;pH 7.4) maintained at 37° C. containing (in mM): 120 NaCl, 25 NaHCO₃,3.3 KH₂PO₄, 0.8 K₂HPO₄, 1.2 CaCl₂, 1.2 MgCl₂, and 10 glucose. Thesolution osmolarity was between 280 and 300 mOsmol/kg H₂O for allphysiological salt solutions used. Cells were voltage clamped to 0 mV(model EVC4000; WPI). RT was measured by applying a 1- or 2-mV pulse at30-s intervals and calculating RT by Ohm's law. Data were recorded usinga PowerLab workstation (ADInstruments).

Test compounds were prepared as a 10 mM stock solution in DMSO (95%).Serial 3-fold dilutions were freshly prepared in an appropriate vehicle(distilled H₂O or Ringers solution). The initial concentration was addedto the apical chamber as a 1000× concentrate in 5 μL, resulting in afinal 1× concentration the 5 mL volume of the Ussing chamber. Subsequentadditions of compound were added in a 3.3 μL volume of the 1000×serially diluted stock solution. At the completion of theconcentration-response experiment, amiloride (10 μM) was added into theapical chamber to enable the total amiloride-sensitive current to bemeasured. An amiloride control IC₅₀ was established at the start of eachexperiment.

Results are expressed as the mean % inhibition of theamiloride-sensitive ISC. Concentration-response curves were plotted andIC₅₀ values generated using GraphPad Prism 3.02 or Graphpad Prism 4.Cell inserts were typically run in duplicate and the IC₅₀ calculated onthe mean % inhibition data.

The compounds of the Examples herein below generally have IC₅₀ valuesbelow 10 μM, typically below 1 μM. For instance, the following Exampleshave the stated IC₅₀ values.

HBEC-ENaC/IC50 Example [umol I-1] 1.0 0.017 1.1 0.419 1.2 0.010 1.30.003 1.4 0.302 2.0 0.032 2.1 0.030 2.2 0.012 2.3 0.089 2.4 0.110 3.00.011 4.0 0.101 4.1 0.132 4.2 0.45 5.0 0.012 5.1 0.031 5.2 0.019

The agents of the invention may be administered by any appropriateroute, e.g. orally, for example in the form of a tablet or capsule;parenterally, for example intravenously; topically to the skin;intranasally, for example in the treatment of allergic rhinitis; or,preferably, by inhalation, particularly in the treatment of obstructiveor inflammatory airways diseases. In particular, the agents of theinvention may be delivered as an inhalable formulation for the treatmentof COPD, cystic fibrosis or asthma.

In another aspect, the present invention provides a pharmaceuticalcomposition comprising a compound of the present invention and apharmaceutically acceptable carrier.

The pharmaceutical composition can be formulated for particular routesof administration such as oral administration, parenteraladministration, and rectal administration, etc. In addition, thepharmaceutical compositions of the present invention can be made up in asolid form (including without limitation capsules, tablets, pills,granules, powders or suppositories), or in a liquid form (includingwithout limitation solutions, suspensions or emulsions). Thepharmaceutical compositions can be subjected to conventionalpharmaceutical operations such as sterilization and/or can containconventional inert diluents, lubricating agents, or buffering agents, aswell as adjuvants, such as preservatives, stabilizers, wetting agents,emulsifers and buffers, etc.

Typically, the pharmaceutical compositions are tablets or gelatincapsules comprising the active ingredient together with

a) diluents, e.g., lactose, dextrose, sucrose, mannitol, sorbitol,cellulose and/or glycine;b) lubricants, e.g., silica, talcum, stearic acid, its magnesium orcalcium salt and/or polyethyleneglycol; for tablets alsoc) binders, e.g., magnesium aluminum silicate, starch paste, gelatin,tragacanth, methylcellulose, sodium carboxymethylcellulose and/orpolyvinylpyrrolidone; if desiredd) disintegrants, e.g., starches, agar, alginic acid or its sodium salt,or effervescent mixtures; and/ore) absorbents, colorants, flavors and sweeteners.

Tablets may be either film coated or enteric coated according to methodsknown in the art.

Suitable compositions for oral administration include an effectiveamount of a compound of the invention in the form of tablets, lozenges,aqueous or oily suspensions, dispersible powders or granules, emulsion,hard or soft capsules, or syrups or elixirs. Compositions intended fororal use are prepared according to any method known in the art for themanufacture of pharmaceutical compositions and such compositions cancontain one or more agents selected from the group consisting ofsweetening agents, flavoring agents, coloring agents and preservingagents in order to provide pharmaceutically elegant and palatablepreparations. Tablets may contain the active ingredient in admixturewith nontoxic pharmaceutically acceptable excipients which are suitablefor the manufacture of tablets. These excipients are, for example, inertdiluents, such as calcium carbonate, sodium carbonate, lactose, calciumphosphate or sodium phosphate; granulating and disintegrating agents,for example, corn starch, or alginic acid; binding agents, for example,starch, gelatin or acacia; and lubricating agents, for example magnesiumstearate, stearic acid or talc. The tablets are uncoated or coated byknown techniques to delay disintegration and absorption in thegastrointestinal tract and thereby provide a sustained action over alonger period. For example, a time delay material such as glycerylmonostearate or glyceryl distearate can be employed. Formulations fororal use can be presented as hard gelatin capsules wherein the activeingredient is mixed with an inert solid diluent, for example, calciumcarbonate, calcium phosphate or kaolin, or as soft gelatin capsuleswherein the active ingredient is mixed with water or an oil medium, forexample, peanut oil, liquid paraffin or olive oil.

Certain injectable compositions are aqueous isotonic solutions orsuspensions, and suppositories are advantageously prepared from fattyemulsions or suspensions. Said compositions may be sterilized and/orcontain adjuvants, such as preserving, stabilizing, wetting oremulsifying agents, solution promoters, salts for regulating the osmoticpressure and/or buffers. In addition, they may also contain othertherapeutically valuable substances. Said compositions are preparedaccording to conventional mixing, granulating or coating methods,respectively, and contain about 0.1-75%, or contain about 1-50%, of theactive ingredient.

Suitable compositions for transdermal application include an effectiveamount of a compound of the invention with a suitable carrier. Carrierssuitable for transdermal delivery include absorbable pharmacologicallyacceptable solvents to assist passage through the skin of the host. Forexample, transdermal devices are in the form of a bandage comprising abacking member, a reservoir containing the compound optionally withcarriers, optionally a rate controlling barrier to deliver the compoundof the skin of the host at a controlled and predetermined rate over aprolonged period of time, and means to secure the device to the skin.

Suitable compositions for topical application, e.g., to the skin andeyes, include aqueous solutions, suspensions, ointments, creams, gels orsprayable formulations, e.g., for delivery by aerosol or the like. Suchtopical delivery systems will in particular be appropriate for dermalapplication, e.g., for the treatment of skin cancer, e.g., forprophylactic use in sun creams, lotions, sprays and the like. They arethus particularly suited for use in topical, including cosmetic,formulations well-known in the art. Such may contain solubilizers,stabilizers, tonicity enhancing agents, buffers and preservatives.

As used herein a topical application may also pertain to an inhalationor to an intranasal application. They may be conveniently delivered inthe form of a dry powder (either alone, as a mixture, for example a dryblend with lactose, or a mixed component particle, for example withphospholipids) from a dry powder inhaler or an aerosol spraypresentation from a pressurised container, pump, spray, atomizer ornebuliser, with or without the use of a suitable propellant.

Where the inhalable form of the active ingredient is an aerosolcomposition, the inhalation device may be an aerosol vial provided witha valve adapted to deliver a metered dose, such as 10 to 100 μl, e.g. 25to 50 μl, of the composition, i.e. a device known as a metered doseinhaler. Suitable such aerosol vials and procedures for containingwithin them aerosol compositions under pressure are well known to thoseskilled in the art of inhalation therapy. For example, an aerosolcomposition may be administered from a coated can, for example asdescribed in EP-A-0642992. Where the inhalable form of the activeingredient is a nebulizable aqueous, organic or aqueous/organicdispersion, the inhalation device may be a known nebulizer, for examplea conventional pneumatic nebulizer such as an airjet nebulizer, or anultrasonic nebulizer, which may contain, for example, from 1 to 50 ml,commonly 1 to 10 ml, of the dispersion; or a hand-held nebulizer,sometimes referred to as a soft mist or soft spray inhaler, for examplean electronically controlled device such as an AERx (Aradigm, US) orAerodose (Aerogen), or a mechanical device such as a RESPIMAT(Boehringer Ingelheim) nebulizer which allows much smaller nebulizedvolumes, e.g. 10 to 100 μl, than conventional nebulizers. Where theinhalable form of the active ingredient is the finely dividedparticulate form, the inhalation device may be, for example, a drypowder inhalation device adapted to deliver dry powder from a capsule orblister containing a dry powder comprising a dosage unit of (A) and/or(B) or a multidose dry powder inhalation (MDPI) device adapted todeliver, for example, 3-25 mg of dry powder comprising a dosage unit of(A) and/or (B) per actuation. The dry powder composition preferablycontains a diluent or carrier, such as lactose, and a compound thathelps to protect against product performance deterioration due tomoisture e.g. magnesium stearate. Suitable such dry powder inhalationdevices include devices disclosed in U.S. Pat. No. 3,991,761 (includingthe AEROLIZER™ device), WO 05/113042 (including the BREEZHALER™ device),WO 97/20589 (including the CERTIHALER™ device), WO 97/30743 (includingthe TWISTHALER™ device), WO 05/37353 (including the GYROHALER™ device),U.S. Pat. No. 6,536,427 (including the DISKUS™ device), WO 97/25086(including the DISKHALER™ device), WO 95/14089 (including the GEMINI™device), WO 03/77979 (including the PROHALER™ device), and also thedevices disclosed in WO 08/51621, WO 09/117112 and US 2005/0183724.

The invention also includes (A) an agent of the invention in free form,or a pharmaceutically acceptable salt or solvate thereof, in inhalableform; (B) an inhalable medicament comprising such a compound ininhalable form together with a pharmaceutically acceptable carrier ininhalable form; (C) a pharmaceutical product comprising such a compoundin inhalable form in association with an inhalation device; and (D) aninhalation device containing such a compound in inhalable form.

Dosages of agents of the invention employed in practising the presentinvention will of course vary depending, for example, on the particularcondition to be treated, the effect desired and the mode ofadministration. In general, suitable daily dosages for administration byinhalation are of the order of 0.0001 to 30 mg/kg, typically 0.01 to 10mg per patient, while for oral administration suitable daily doses areof the order of 0.01 to 100 mg/kg.

The present invention further provides anhydrous pharmaceuticalcompositions and dosage forms comprising the compounds of the presentinvention as active ingredients, since water may facilitate thedegradation of certain compounds.

Anhydrous pharmaceutical compositions and dosage forms of the inventioncan be prepared using anhydrous or low moisture containing ingredientsand low moisture or low humidity conditions. An anhydrous pharmaceuticalcomposition may be prepared and stored such that its anhydrous nature ismaintained. Accordingly, anhydrous compositions are packaged usingmaterials known to prevent exposure to water such that they can beincluded in suitable formulary kits. Examples of suitable packaginginclude, but are not limited to, hermetically sealed foils, plastics,unit dose containers (e.g., vials), blister packs, and strip packs.

The invention further provides pharmaceutical compositions and dosageforms that comprise one or more agents that reduce the rate by which thecompound of the present invention as an active ingredient willdecompose. Such agents, which are referred to herein as “stabilizers,”include, but are not limited to, antioxidants such as ascorbic acid, pHbuffers, or salt buffers, etc.

The compound of the present invention may be administered eithersimultaneously with, or before or after, one or more other therapeuticagent. The compound of the present invention may be administeredseparately, by the same or different route of administration, ortogether in the same pharmaceutical composition as the other agents.

In one embodiment, the invention provides a product comprising acompound of formula (I) and at least one other therapeutic agent as acombined preparation for simultaneous, separate or sequential use intherapy. In one embodiment, the therapy is the treatment of a disease orcondition mediated by blockade of the epithelial sodium channel.Products provided as a combined preparation include a compositioncomprising the compound of formula (I) and the other therapeuticagent(s) together in the same pharmaceutical composition, or thecompound of formula (I) and the other therapeutic agent(s) in separateform, e.g. in the form of a kit.

In one embodiment, the invention provides a pharmaceutical compositioncomprising a compound of formula (I) and another therapeutic agent(s).Optionally, the pharmaceutical composition may comprise apharmaceutically acceptable excipient, as described above.

In one embodiment, the invention provides a kit comprising two or moreseparate pharmaceutical compositions, at least one of which contains acompound of formula (I). In one embodiment, the kit comprises means forseparately retaining said compositions, such as a container, dividedbottle, or divided foil packet. An example of such a kit is a blisterpack, as typically used for the packaging of tablets, capsules and thelike.

The kit of the invention may be used for administering different dosageforms, for example, oral and parenteral, for administering the separatecompositions at different dosage intervals, or for titrating theseparate compositions against one another. To assist compliance, the kitof the invention typically comprises directions for administration.

The compounds of, formula (I) and their pharmaceutically acceptablesalts and solvates have the advantage that they are more selective, havea more rapid onset of action, are more potent, are better absorbed, aremore stable, are more resistant to metabolism, have a reduced ‘foodeffect’, have an improved safety profile or have other more desirableproperties (e.g. with respect to solubility or hygroscopicity) than thecompounds of the prior art.

In particular, the compounds of the invention exhibit an advantageousstability profile in human plasma. In providing compounds which exhibitan advantageous stability profile in human plasma, the inventionprovides compounds which effectively blockade the epithelial sodiumchannel (ENaC) with improved pharmacokinetics.

The stability of the compounds of the invention in human plasma may bemeasured as follows:

Stability in Human Plasma

Blood was taken from healthy volunteers in lithium heparin tubes andplasma was prepared by centrifugal separation of blood cells at 1500 g.Plasma from at least 3 individuals was pooled and used for compoundstability determination.

Compounds were prepared in 100% DMSO at a concentration of 10 mM andserially diluted to a concentration of 100 μM. Incubations wereperformed at a final concentration of 1 μM and were initiated byaddition of 3 μl of 100 μM compound stock solution into 297 μl plasma,pre-warmed in a water bath to 37° C., followed by brief vortex mixing.50 μl aliquots of plasma were removed at 4 timepoints over a 1 hourincubation period and quenched immediately by protein precipitationachieved by addition of the aliquot of plasma to a pre-prepared 96 wellplate containing 150 μl of acetonitrile containing an appropriateinternal standard. Acetonitrile quenched samples were vortex mixedfollowed by centrifugation for 20 minutes at 1500 g to remove theprecipitated proteins. Initial compound levels (at time=0 minutes) wasestablished by spiking compound into pre-quenched plasma at the sameconcentration as used in the incubation. The supernatant was thenremoved, diluted 1:1 with water and analysed for compound remaining byliquid chromatography-tandem mass spectrometry (Waters Acquity UPLC,Applied Biosystems API4000). The plasma elimination rate constant of thecompound was calculated by fitting the peak area ratio of compound:internal standard to an exponential decay function and the half lifecalculated by division of the natural log of 2 by the elimination rateconstant (Microsoft Excel).

The following Examples have the stated half life in human plasma values.

Half life in human Example plasma (mins) 1.0 2 1.2 3 1.3 4 2.0 1 2.1 53.0 2 4.0 28 4.1 3 4.2 4 5.0 5 5.1 2 5.2 39

In the combination therapies of the invention, the compound of theinvention and the other therapeutic agent may be manufactured and/orformulated by the same or different manufacturers. Moreover, thecompound of the invention and the other therapeutic may be broughttogether into a combination therapy: (i) prior to release of thecombination product to physicians (e.g. in the case of a kit comprisingthe compound of the invention and the other therapeutic agent); (ii) bythe physician themselves (or under the guidance of the physician)shortly before administration; (iii) in the patient themselves, e.g.during sequential administration of the compound of the invention andthe other therapeutic agent.

Accordingly, the invention provides the use of a compound of formula (I)for treating a disease or condition mediated by blockade of theepithelial sodium channel, wherein the medicament is prepared foradministration with another therapeutic agent. The invention alsoprovides the use of another therapeutic agent for treating a disease orcondition mediated by blockade of the epithelial sodium channel, whereinthe medicament is administered with a compound of formula (I).

The invention also provides a compound of formula (I) for use in amethod of treating a disease or condition mediated by blockade of theepithelial sodium channel, wherein the compound of formula (I) isprepared for administration with another therapeutic agent. Theinvention also provides another therapeutic agent for use in a method oftreating a disease or condition mediated by blockade of the epithelialsodium channel, wherein the other therapeutic agent is prepared foradministration with a compound of formula (I). The invention alsoprovides a compound of formula (I) for use in a method of treating adisease or condition mediated by blockade of the epithelial sodiumchannel, wherein the compound of formula (I) is administered withanother therapeutic agent. The invention also provides anothertherapeutic agent for use in a method of treating a disease or conditionmediated by blockade of the epithelial sodium channel, wherein the othertherapeutic agent is administered with a compound of formula (I).

The invention also provides the use of a compound of formula (I) fortreating a disease or condition mediated by blockade of the epithelialsodium channel, wherein the patient has previously (e.g. within 24hours) been treated with another therapeutic agent. The invention alsoprovides the use of another therapeutic agent for treating a disease orcondition mediated by blockade of the epithelial sodium channel, whereinthe patient has previously (e.g. within 24 hours) been treated with acompound of formula (I).

In one embodiment, the other therapeutic agent is selected fromanti-inflammatory, bronchodilatory, antihistamine, decongestant andanti-tussive drug substances, particularly in the treatment of cysticfibrosis or obstructive or inflammatory airways diseases such as thosementioned hereinbefore, e.g., as potentiators of therapeutic activity ofsuch drugs or as a means of reducing required dosaging or potential sideeffects of such drugs.

Accordingly, the invention includes as a further aspect a combination ofan epithelial sodium channel blocker of the present invention withosmotic agents (hypertonic saline, dextran, mannitol, Xylitol),modifiers of CFTR function, both wild-type and mutant (correctors andpotentiators), e.g., those described in WO 2007/021982, WO 2006/099256,WO 2006/127588, WO 2004/080972, WO 2005/026137, WO 2005/035514, WO2005/075435, WO 2004/111014, WO 2006/101740, WO 2004/110352, WO2005/120497 and US 2005/0176761, an anti-inflammatory, bronchodilatory,antihistamine, anti-tussive, antibiotic or DNase drug substance, saidepithelial sodium channel blocker and said drug substance being in thesame or different pharmaceutical composition.

Suitable modifiers of CFTR function include CFTR potentiators, inparticular the compound VX-770 of formula

Suitable antibiotics include macrolide antibiotics, e.g., tobramycin(TOBI™).

Suitable DNase drug substances include dornase alfa (Pulmozyme™), ahighly-purified solution of recombinant human deoxyribonuclease I(rhDNase), which selectively cleaves DNA. Dornase alfa is used to treatcystic fibrosis.

Other useful combinations of epithelial sodium channel blockers withanti-inflammatory drugs are those with antagonists of chemokinereceptors, e.g., CCR-1, CCR-2, CCR-3, CCR-4, CCR-5, CCR-6, CCR-7, CCR-8,CCR-9 and CCR10, CXCR1, CXCR2, CXCR3, CXCR4, CXCR5, particularly CCR-5antagonists, such as Schering-Plough antagonists SC-351125, SCH-55700and SCH-D; Takeda antagonists, such asN-[[4-[[[6,7-dihydro-2-(4-methyl-phenyl)-5H-benzo-cyclohepten-8-yl]carbonyl]amino]phenyl]-methyl]tetrahydro-N,N-dimethyl-2H-pyran-4-amin-iumchloride (TAK-770); and CCR-5 antagonists described in U.S. Pat. No.6,166,037 (particularly claims 18 and 19), WO 00/66558 (particularlyclaim 8), WO 00/66559 (particularly claim 9), WO 04/018425 and WO04/026873.

Suitable anti-inflammatory drugs include steroids, in particular,glucocorticosteroids, such as budesonide, beclamethasone dipropionate,fluticasone propionate, ciclesonide or mometasone furoate, or steroidsdescribed in WO 02/88167, WO 02/12266, WO 02/100879, WO 02/00679(especially those of Examples 3, 11, 14, 17, 19, 26, 34, 37, 39, 51, 60,67, 72, 73, 90, 99 and 101), WO 03/35668, WO 03/48181, WO 03/62259, WO03/64445, WO 03/72592, WO 04/39827 and WO 04/66920; non-steroidalglucocorticoid receptor agonists, such as those described in DE10261874, WO 00/00531, WO 02/10143, WO 03/82280, WO 03/82787, WO03/86294, WO 03/104195, WO 03/101932, WO 04/05229, WO 04/18429, WO04/19935 and WO 04/26248; LTD4 antagonists, such as montelukast andzafirlukast; PDE4 inhibitors, such as cilomilast (Ariflo®GlaxoSmithKline), Roflumilast (Byk Gulden), V-11294A (Napp), BAY19-8004(Bayer), SCH-351591 (Schering-Plough), Arofylline (AlmirallProdesfarma), PD189659/PD168787 (Parke-Davis), AWD-12-281 (Asta Medica),CDC-801 (Celgene), SelCID™ CC-10004 (Celgene), VM554/UM565 (Vernalis),T-440 (Tanabe), KW-4490 (Kyowa Hakko Kogyo), and those disclosed in WO92/19594, WO 93/19749, WO 93/19750, WO 93/19751, WO 98/18796, WO99/16766, WO 01/13953, WO 03/104204, WO 03/104205, WO 03/39544, WO04/000814, WO 04/000839, WO 04/005258, WO 04/018450, WO 04/018451, WO04/018457, WO 04/018465, WO 04/018431, WO 04/018449, WO 04/018450, WO04/018451, WO 04/018457, WO 04/018465, WO 04/019944, WO 04/019945, WO04/045607 and WO 04/037805; adenosine A2B receptor antagonists such asthose described in WO 02/42298; and beta-2 adrenoceptor agonists, suchas albuterol (salbutamol), metaproterenol, terbutaline, salmeterolfenoterol, procaterol, and especially, formoterol, carmoterol andpharmaceutically acceptable salts thereof, and compounds (in free orsalt or solvate form) of formula (I) of WO 0075114, which document isincorporated herein by reference, preferably compounds of the Examplesthereof, especially a compound of formula:

corresponding to indacaterol and pharmaceutically acceptable saltsthereof, as well as compounds (in free or salt or solvate form) offormula (I) of WO 04/16601, and also compounds of EP 1440966, JP05025045, WO 93/18007, WO 99/64035, USP 2002/0055651, WO 01/42193, WO01/83462, WO 02/66422, WO 02/70490, WO 02/76933, WO 03/24439, WO03/42160, WO 03/42164, WO 03/72539, WO 03/91204, WO 03/99764, WO04/16578, WO 04/22547, WO 04/32921, WO 04/33412, WO 04/37768, WO04/37773, WO 04/37807, WO 04/39762, WO 04/39766, WO 04/45618, WO04/46083, WO 04/80964, WO 04/108765 and WO 04/108676.

Suitable bronchodilatory drugs include anticholinergic or antimuscarinicagents, in particular, ipratropium bromide, oxitropium bromide,tiotropium salts and CHF 4226 (Chiesi), and glycopyrrolate, but alsothose described in EP 424021, U.S. Pat. No. 3,714,357, U.S. Pat. No.5,171,744, WO 01/04118, WO 02/00652, WO 02/51841, WO 02/53564, WO03/00840, WO 03/33495, WO 03/53966, WO 03/87094, WO 04/018422 and WO04/05285.

Suitable dual anti-inflammatory and bronchodilatory drugs include dualbeta-2 adrenoceptor agonist/muscarinic antagonists such as thosedisclosed in USP 2004/0167167, WO 04/74246 and WO 04/74812.

Suitable antihistamine drug substances include cetirizine hydrochloride,acetaminophen, clemastine fumarate, promethazine, loratidine,desloratidine, diphenhydramine and fexofenadine hydrochloride,activastine, astemizole, azelastine, ebastine, epinastine, mizolastineand tefenadine, as well as those disclosed in JP 2004107299, WO03/099807 and WO 04/026841.

As used herein, the term “pharmaceutically acceptable carrier” includesany and all solvents, dispersion media, coatings, surfactants,antioxidants, preservatives (e.g., antibacterial agents, antifungalagents), isotonic agents, absorption delaying agents, salts,preservatives, drugs, drug stabilizers, binders, excipients,disintegration agents, lubricants, sweetening agents, flavoring agents,dyes, and the like and combinations thereof, as would be known to thoseskilled in the art (see, for example, Remington's PharmaceuticalSciences, 18th Ed. Mack Printing Company, 1990, pp. 1289-1329). Exceptinsofar as any conventional carrier is incompatible with the activeingredient, its use in the therapeutic or pharmaceutical compositions iscontemplated.

The term “a therapeutically effective amount” of a compound of thepresent invention refers to an amount of the compound of the presentinvention that will elicit the biological or medical response of asubject, for example, reduction or inhibition of an enzyme or a proteinactivity, or ameliorate symptoms, alleviate conditions, slow or delaydisease progression, or prevent a disease, etc. In one non-limitingembodiment, the term “a therapeutically effective amount” refers to theamount of the compound of the present invention that, when administeredto a subject, is effective to (1) at least partially alleviating,inhibiting, preventing and/or ameliorating a condition, or a disorder ora disease (i) mediated by the epithelial sodium channel, or (ii)associated with epithelial sodium channel activity, or (iii)characterized by activity (normal or abnormal) of the epithelial sodiumchannel; or (2) reducing or inhibiting the activity of the epithelialsodium channel. In another non-limiting embodiment, the term “atherapeutically effective amount” refers to the amount of the compoundof the present invention that, when administered to a cell, or a tissue,or a non-cellular biological material, or a medium, is effective to atleast partially reducing or inhibiting the activity of the epithelialsodium channel.

As used herein, the term “subject” refers to an animal. Typically theanimal is a mammal. A subject also refers to for example, primates(e.g., humans), cows, sheep, goats, horses, dogs, cats, rabbits, rats,mice, fish, birds and the like. In certain embodiments, the subject is aprimate. In yet other embodiments, the subject is a human.

As used herein, the term “inhibit”, “inhibition” or “inhibiting” refersto the reduction or suppression of a given condition, symptom, ordisorder, or disease, or a significant decrease in the baseline activityof a biological activity or process.

As used herein, the term “treat”, “treating” or “treatment” of anydisease or disorder refers in one embodiment, to ameliorating thedisease or disorder (i.e., slowing or arresting or reducing thedevelopment of the disease or at least one of the clinical symptomsthereof). In another embodiment “treat”, “treating” or “treatment”refers to alleviating or ameliorating at least one physical parameterincluding those which may not be discernible by the patient. In yetanother embodiment, “treat”, “treating” or “treatment” refers tomodulating the disease or disorder, either physically, (e.g.,stabilization of a discernible symptom), physiologically, (e.g.,stabilization of a physical parameter), or both. In yet anotherembodiment, “treat”, “treating” or “treatment” refers to preventing ordelaying the onset or development or progression of the disease ordisorder.

As used herein, a subject is “in need of” a treatment if such subjectwould benefit biologically, medically or in quality of life from suchtreatment.

As used herein, the term “a,” “an,” “the” and similar terms used in thecontext of the present invention (especially in the context of theclaims) are to be construed to cover both the singular and plural unlessotherwise indicated herein or clearly contradicted by the context.

All methods described herein can be performed in any suitable orderunless otherwise indicated herein or otherwise clearly contradicted bycontext. The use of any and all examples, or exemplary language (e.g.“such as”) provided herein is intended merely to better illuminate theinvention and does not pose a limitation on the scope of the inventionotherwise claimed.

The invention is illustrated by the following Examples.

EXAMPLES

Referring to the examples that follow, compounds of the preferredembodiments are synthesized using the methods described herein, or othermethods, which are known in the art.

It should be understood that the organic compounds according to thepreferred embodiments may exhibit the phenomenon of tautomerism. As thechemical structures within this specification can only represent one ofthe possible tautomeric forms, it should be understood that thepreferred embodiments encompasses any tautomeric form of the drawnstructure.

It is understood that the invention is not limited to the embodimentsset forth herein for illustration, but embraces all such forms thereofas come within the scope of the above disclosure.

General Conditions:

Mass spectra were run on LCMS systems using electrospray ionization.These were either Agilent 1100 HPLC/Micromass Platform Mass Spectrometercombinations, or Agilent 1200 HPLC/Agilent 6130 Quadropole MassSpectrometer combinations, or Waters Acquity UPLC with SQD MassSpectrometer. [M+H]⁺ refers to mono-isotopic molecular weights.

NMR spectra were run on open access Bruker AVANCE 400 NMR spectrometersusing ICON-NMR. Spectra were measured at 298K and were referenced usingthe solvent peak. Some protons were not observed directly due to thevery broad nature of their exchangeable resonances.

The following examples are intended to illustrate the invention and arenot to be construed as being limitations thereon. Temperatures are givenin degrees centigrade. If not mentioned otherwise, all evaporations areperformed under reduced pressure. The structure of final products,intermediates and starting materials is confirmed by standard analyticalmethods, e.g., microanalysis and spectroscopic characteristics, e.g.,MS, IR, NMR. Abbreviations used are those conventional in the art. Ifnot defined, the terms have their generally accepted meanings.

Abbreviations

Br broadd doubletDCM dichloromethaneDSC differential scanning calorimetry

DMF N,N-dimethylformamide

DMI 1,3-dimethyl-2-imidazolidinoneDMSO dimethylsulfoxideEDCI N-(3-dimethylaminopropyl)-N′-ethylcarbodiimide

EtOAc EtOAc

h hour(s)HOBt 1-hydroxybenzotriazoleHPLC high pressure liquid chromatographyLC-MS liquid chromatography and mass spectrometryMeOH methanolMS mass spectrometrym multiplet2-meTHF 2-methyltetrahydrofuranmin minutesml milliliter(s)m/z mass to charge ratioNMR nuclear magnetic resonanceiPrOH isopropanolppm parts per millionPS polymer supportedPEAX PE-anion exchange (e.g. Isolute® PE-AX columns from Biotage)Rt retention times singletSCX-2 strong cation exchange (e.g. Isolute® SCX-2 columns from Biotage)t tripletTEA triethylamineTFA trifluoroacetic acidTHF tetrahydrofuran

Referring to the examples that follow, compounds of the preferredembodiments were synthesized using the methods described herein, orother methods, which are known in the art.

The various starting materials, intermediates, and compounds of thepreferred embodiments may be isolated and purified, where appropriate,using conventional techniques such as precipitation, filtration,crystallization, evaporation, distillation, and chromatography. Unlessotherwise stated, all starting materials are obtained from commercialsuppliers and used without further purification. Salts may be preparedfrom compounds by known salt-forming procedures.

It should be understood that the organic compounds according to thepreferred embodiments may exhibit the phenomenon of tautomerism. As thechemical structures within this specification can only represent one ofthe possible tautomeric forms, it should be understood that thepreferred embodiments encompasses any tautomeric form of the drawnstructure.

If not indicated otherwise, the analytical HPLC conditions are asfollows:

Method 2minLC_v002

-   Column Waters BEH C18 50×2.1 mm, 1.7 μm-   Column Temperature 50° C.-   Eluents A: H₂O, B: methanol, both containing 0.1% TFA-   Flow Rate 0.8 ml/min-   Gradient 0.20 min 5% B; 5% to 95% B in 1.30 min, 0.25 min 95% B    Method 2minLC_v003-   Column Waters BEH C18 50×2.1 mm, 1.7 μm-   Column Temperature 50° C.-   Eluents A: H₂O, B: acetonitrile, both containing 0.1% TFA-   Flow Rate 0.8 mL/min-   Gradient 0.20 min 5% B; 5% to 95% B in 1.30 min, 0.25 min 95% B    Method 10minLC_v002-   Column Waters BEH C18 50×2.1 mm, 1.7 μm-   Column Temperature 50° C.-   Eluents A: H₂O, B: methanol, both containing 0.1% TFA-   Flow Rate 0.8 mL/min-   Gradient 0.20 min 5% B; 5% to 95% B in 7.80 min, 1.00 min 95% B    Method 10minLC_v003-   Column Waters BEH C18 50×2.1 mm, 1.7 μm-   Column Temperature 50° C.-   Eluents A: H₂O, B: acetonitrile, both containing 0.1% TFA-   Flow Rate 0.8 mL/min-   Gradient 0.20 min 5% B; 5% to 95% B in 7.80 min, 1.00 min 95% B

Method (i)

-   Column Agilent Zorbax SB-C18 (Rapid resolution) 30×2.1 mm, 3.5 μm-   Column Temperature 30° C.-   Eluents B: H₂O, C: acetonitrile, both containing 0.1% formic acid-   Flow Rate 0.8 mL/min-   Gradient 1 min 5% C; 5% to 95% C in 5 min, 3.00 min 95% C

Method (ii)

-   Column SB-C18 50×4.6 mm, 1.8 μM-   Column Temperature 30° C.-   Eluents A: H₂O, B: acetonitrile, both containing 0.1% formic acid-   Flow Rate 1 mL/min-   Gradient 1 min 2% B; 2% to 70% B in 4 min, 70% to 90% B in 0.1 min,    4.9 min 95% B

Example compounds of the present invention include:

Preparation of Final Compounds Example 1.03-(3-{2-[(E)-3,5-Diamino-6-chloro-pyrazine-2-carbonylimino]-1,3,8-triaza-spiro[4.5]decane-8-carbonyl}-benzenesulfonylamino)-propionicacid dipropylcarbamoylmethyl ester succinate salt

Step 1:3-(3-{2-[(E)-3,5-Diamino-6-chloro-pyrazine-2-carbonylimino]-1,3,8-triaza-spiro[4.5]decane-8-carbonyl}-benzenesulfonylamino)-propionicacid dipropylcarbamoylmethyl ester

The title compound may be prepared by either Method A or B

Method A:

To a stirred solution of3-(2-dipropylcarbamoylmethoxycarbonyl-ethylsulfamoyl)-benzoic acid (Int.AA) (7.0 g, 16.89 mmol) in DMF at RT was added3,5-diamino-6-chloro-pyrazine-2-carboxylic acid[1,3,8-triaza-spiro[4.5]dec-(2E)-ylidene]-amide hydrochloride(WO09074575, Ex. 38, page 123) (8.20 g, 16.89 mmol) followed byN-methylmorpholine (7.43 ml, 67.6 mmol). The reaction mixture wasstirred at RT for 10 min and treated with HATU (6.42 g, 16.89 mmol) inone portion. The mixture was stirred at RT for a further 10 min and thenthe reaction was quenched by addition of ice-water (500 ml). Theresulting solid was collected by filtration and dissolved in DCM. Thesolution was washed with water (1×500 ml), dried (MgSO₄) andconcentrated in vacuo to give a crude orange oil. Purification bychromatography on silica eluting with DCM/iPA (2% TEA) afforded thetitle compound as the hexafluorophosphate salt; LC-MS Rt 3.72 mins;721.5 [M+H]⁺, Method 10minLC_v003.; ¹H NMR (400 MHz, DMSO-d6) δ 8.52(1H, br s), 8.35 (1H, br s), 7.88 (2H, dt), 7.77 (1H, s), 7.72-7.65 (2H,m), 6.72 (2H, br s), 4.73 (2H, s), 3.80 (1H, b s), 3.61 (1H, b s), 3.43(3H, b s), 3.17-3.09 (4H, m), 3.04 (2H, t), 2.53 (2H, under DMSO), 1.78(2H, b s), 1.69 (2H, b s), 1.56-1.38 (4H, m), 0.84 (3H, t), 0.78 (3H, t)

Method B:

To a stirred solution of3-(2-dipropylcarbamoylmethoxycarbonyl-ethylsulfamoyl)-benzoic acid (Int.AA) (6.1 g, 12.60 mmol) in THF (50 ml) was added sequentially water (25ml), N-methylmorpholine (7 ml, 63 mmol) and HOBt hydrate (2.9 g, 18.9mmol). The internal temperature was maintained at 20° C.3,5-Diamino-6-chloro-pyrazine-2-carboxylic acid[1,3,8-triaza-spiro[4.5]dec-(2E)-ylidene]-amide hydrochloride(WO09074575, Ex. 38, page 123) (65% purity, 6.3 g, 12.6 mmol) was addedand stirred until a clear solution had formed. EDCI.HCl (3.6 g, 18.9mmol) was added, and the reaction was stirred at RT for 24 h. 2-MeTHF(200 ml) and 2% aq. Na₂CO₃ (150 ml) were added to the reaction mixture.The layers were separated, and the aqueous phase washed with additional2-MeTHF (100 ml). The combined organic layers were washed with 2% aq.Na₂CO₃ (200 ml) and water (2×200 ml). Acetonitrile (100 ml) was added,and the solution concentrated at 30° C. to a volume of 70 ml.Acetonitrile (300 ml) was added, and the solution concentrated again at30° C. to a volume of 150 ml. The solution was heated to 50° C. andmaleic acid (1.62 g) was added to the resulting solution. An off-whiteprecipitate formed immediately, the temperature was allowed to cool toRT over 1 h. The solid was collected by filtration to afford3-(3-{2-[(E)-3,5-Diamino-6-chloro-pyrazine-2-carbonylimino]-1,3,8-triaza-spiro[4.5]decane-8-carbonyl}-benzenesulfonylamino)-propionicacid dipropylcarbamoylmethyl ester as a maleate salt; DCM (200 ml) and2% aq. Na₂CO₃ (200 ml) were added and stirred until the solid had fullydissolved. The organic layer was separated, washed with water (2×100 ml)and concentrated in vacuo to afford the title compound. LC-MS 722.1[M+H]⁺, Method (i).; ¹H NMR (400 MHz, DMSO-d6) δ 9.12-7.57 (4H, br),7.88 (1H, m), 7.77 (1H, m), 7.70 (1H, m), 7.68 (1H, m), 7.05-6.50 (2H,br s), 6.95-6.20 (1H, br s), 4.73 (2H, s), 3.81-3.39 (2H, m), 3.61-3.31(2H, m), 3.43 (2H, br s), 3.15-3.11 (4H, m), 3.04 (2H, t), 2.51 (2H, t),1.79-1.69 (m, 4H), 1.51-1.43 (4H, m), 0.84 (3H, t), 0.78 (3H, t)

Step 2:3-(3-{2-[(E)-3,5-Diamino-6-chloro-pyrazine-2-carbonylimino]-1,3,8-triaza-spiro[4.5]decane-8-carbonyl}-benzenesulfonylamino)-propionicacid dipropylcarbamoylmethyl ester succinate salt

The title compound may be prepared by either Method C or D

Method C:

Succinic acid (409 mg, 3.47 mmol) was added to a solution of3-(3-{2-[(E)-3,5-diamino-6-chloro-pyrazine-2-carbonylimino]-1,3,8-triaza-spiro[4.5]decane-8-carbonyl}-benzenesulfonylamino)-propionicacid dipropylcarbamoylmethyl ester (step 1, Method B)(2.50 g, 3.47 mmol)in acetonitrile (25 ml) and water (1.5 ml) at 50° C. The resulting clearsolution was cooled to RT over 30 mins. Crystallisation started to occurat an internal temperature of ˜30° C. The resulting slurry was stirredfor at RT for 16 h, The crystals were collected by filtration, and thefilter cake washed with acetonitrile/water (95:5) and dried at 50° C.under vacuum to afford the title compound;

Method D:

A mixture comprising succinic acid (0.50 g, 4.23 mmol) and acetone (20g) was heated to 45° C. until a clear solution formed and then filtered(0.2 μm PTFE filter). In a second reaction vessel,3-(3-{2-[(E)-3,5-diamino-6-chloro-pyrazine-2-carbonylimino]-1,3,8-triaza-spiro[4.5]decane-8-carbonyl}-benzenesulfonylamino)-propionicacid dipropylcarbamoylmethyl ester (step 1, Method B)(3.00 g, 4.16 mmol)and acetone (30 g) were heated to 45° C. until a clear solution formedand then filtered (0.2 μm PTFE filter).

The solution of succinic acid (0.50 g, 4.23 mmol) in acetone (20 ml) washeated at 45° C. for 1 h and treated with a portion of the solution of3-(3-{2-[(E)-3,5-diamino-6-chloro-pyrazine-2-carbonylimino]-1,3,8-triaza-spiro[4.5]decane-8-carbonyl}-benzenesulfonylamino)-propionic acid dipropylcarbamoylmethyl ester in acetone (1.62 gof solution) over 10 min. The resulting mixture was treated with asuspension of seed crystals of3-(3-{2-[(E)-3,5-diamino-6-chloro-pyrazine-2-carbonylimino]-1,3,8-triaza-spiro[4.5]decane-8-carbonyl}-benzenesulfonylamino)-propionicacid dipropylcarbamoylmethyl ester (as prepared using method C, 20 mg)in acetone (300 mg) and stirred at 45° C. for 30 min. The remainingsolution of3-(3-{2-[(E)-3,5-diamino-6-chloro-pyrazine-2-carbonylimino]-1,3,8-triaza-spiro[4.5]decane-8-carbonyl}-benzenesulfonylamino)-propionic acid dipropylcarbamoyl methyl ester in acetone (31.38g) was added to the mixture over 5 h and heating continued at 45° C. for1 h. The suspension was cooled to 25° C. over 1 h and stirred for afurther 1 h. The suspension was filtered over a glass frit and thefilter cake was washed with acetone (2×5 g). The filter cake was driedat 50° C. to afford the title compound; HPLC Rt 4.02 min, method ii; ¹HNMR (400 MHz, DMSO-d6) δ 7.87 (1H, m), 7.78 (1H, m), 7.69 (1H, m), 7.68(1H, m), 6.85 (2H, br s), 4.73 (2H, s), 3.84-3.20 (6H and water, brhump), 3.17-3.09 (4H, m), 3.04 (2H, t), 2.53 (2H, under DMSO), 2.39 (4H,s), 1.80 (2H, br s), 1.70 (2H, br s), 1.55-1.37 (4H, m), 0.85 (3H, t),0.78 (3H, t) (Please note: The two exchangeable succinate protons and3-4 acidic NH resonances were not observed directly due to the verybroad nature of some of the exchangeable resonances; Melting TemperatureT_(m) (DSC)=149° C.

The compounds of the following tabulated Examples (Table 1) wereprepared by a similar method to that of Example 1.0 replacing3-(2-dipropylcarbamoylmethoxycarbonyl-ethylsulfamoyl)-benzoic acid (Int.AA) with the appropriate Intermediate (the preparations of which aredescribed hereinafter or are commercially available).

TABLE 1 Ex. Name [M + H]⁺/NMR 1.1 [4-(3-{2-[(Z)-3,5-Diamino-6- ¹H NMR(400 MHz, DMSO-d6) δ 8.67 (1H, s), chloro-pyrazine-2- 8.44 (1H, m), 7.45(1H, m), 7.16 (2H, d), 6.88 carbonylimino]-1,3,8-triaza- (2H, d), 5.76(1H, s),4.97 (1H, s), 4.89 (1H, m). spiro[4.5]dec-8-yl}-3-oxo-propyl)-4.81 (2H, d), 3.75 (1H, m), 3.68-3.4 (6H, m), phenoxy]-acetic acid [(2-2.96 (1H, s), 2.83 (2H, s), 2.75 (2H, t), 2.61 (2H,hydroxy-ethyl)-methyl- t), 1.67 (4H, m). LC-MS Rt 1.04 mins;carbamoyl]-methyl ester 646.5 [M + H]⁺, Method 2minLC_v002. 1.2[4-(3-{2-[(E)-3,5-Diamino-6- ¹H NMR (400 MHz, DMSO-d6) δ 8.98-8.27 (2H,chloro-pyrazine-2- b hump), 7.43-6.40 (NH2, b hump), 7.17 (2H, d),carbonylimino]-1,3,8-triaza- 6.86 (2H, d), 4.85 (2H, s), 4.77-4.71 (3H,m), spiro[4.5]dec-8-yl}-3-oxo-propyl)- 3.76-3.65 (1H, m), 3.62-3.57 (1H,m), 3.50-3.41 phenoxy]-acetic acid cyclohexyl (2H, m), 3.39-3.33 (1H,m), 2.75 (2H, t), 2.59 oxycarbonylmethyl ester (2H, t), 1.79-1.74 (2H,m), 1.71-1.59 (6H, m), 1.51-1.19 (7H, m). LC-MS Rt 1.31 mins;671.3/673.3 [M + H]+, Method 2minLC_v002. 1.33-(3-{2-[(E)-3,5-Diamino-6- 1H NMR (400 MHz, DMSO-d6) δ 8.52 (NH, b s),chloro-pyrazine-2- 8.35 (NH, b s), 7.88 (2H, dt), 7.77 (1H, s),7.72-7.66 carbonylimino]-1,3,8-triaza- (2H, m), 6.71 (NH2, b s),4.73-4.67 (1H, m), spiro[4.5]decane-8-carbonyl}- 4.59 (2H, s), 3.79 (1H,b s), 3.61 (1H, b s), 3.43 benzene sulfonylamino)- (3H, b s), 3.02 (2H,t), 2.55 (2H, t), 1.82-1.57 (8H, propionic acid cyclohexyloxy m),1.47-1.17 (6H, m). LC-MS Rt 3.99 mins; carbonylmethyl ester 720.5 [M +H]⁺, Method 10minLC_v003. 1.4 [4-(3-{2-[(E)-3,5-Diamino-6- 1H NMR (400MHz, DMSO-d6) δ 8.45 (2H, b s), chloro-pyrazine-2- 7.16 (2H, d), 6.88(2H, d), 6.77 (NH2, b s), 4.80 carbonylimino]-1,3,8-triaza- (2H, s),4.01 (2H, s), 3.71-3.62 (1H, m), 3.61-3.52spiro[4.5]dec-8-yl}-3-oxo-propyl)- (1H, m), 3.45-3.35 (4H, m), 2.917(3H, s), 2.82 phenoxy]-acetic acid (3H, s), 2.75 (2H, t), 2.59 (2H, t),1.68-1.54 (4H, dimethylcarbamoylmethyl ester m). LC-MS Rt 3.07 mins;616.4/618.4 [M + H]⁺, Method 10minLC_v002.

TABLE 1.1 Ex. Structure 1.1

1.2

1.3

1.4

Example 2.0[4-(3-{2-[(E)-3,5-Diamino-6-chloro-pyrazine-2-carbonylimino]-1,3,8-triaza-spiro[4.5]dec-8-yl}-3-oxo-propyl)-phenoxy]-aceticacid dipropylcarbamoylmethyl ester

The title compound may be prepared by either Method A or B:

Method A

To a suspension of[4-(3-{2-[(E)-3,5-Diamino-6-chloro-pyrazine-2-carbonylimino]-1,3,8-triaza-spiro[4.5]dec-8-yl}-3-oxo-propyl)-phenoxy]-aceticacid tert-butyl ester, (WO09074575, Ex. 71, page 134) (6.0 g, 8.2 mmol)in 2-MeTHF/Water (60 ml/10 ml) was added NaOH (1.0 g, 25 mmol)portionwise. The reaction mixture was stirred at RT for 2 h. The organiclayer was separated and washed with water (3×10 ml). Water (60 ml) wasadded and the 2-MeTHF was removed in vacuo. THF (30 ml) was added,followed by NaOH (0.68 g, 17 mmol). The reaction mixture was stirred atRT overnight. The organic solvent was removed in vacuo, and theremaining aqueous phase was washed with MTBE (2×30 ml). DMF (30 ml) wasadded, and the pH adjusted to 7 with 4N HCl (4.25 ml) at RT. PhMeazotropical distillation was performed three times. NaHCO₃ (1.0 g, 11.9mmol) and 2-chloro-N,N-dipropyl-acetamide (2.2 g, 12.4 mmol) were addedto the remaining solution, and the resulting reaction mixture was heatedat 60° C. overnight. The mixture was partitioned between DCM (60 ml) andwater (30 ml). The organic phase was washed with water/sat. aq. NaHCO₃(10:1, 5×30 ml), water (30 ml) and concentrated in vacuo to a volume of˜20 ml. 5 ml of the DCM solution was added to isopropyl acetate (20 ml)while stirring at RT. The DCM was removed in vacuo. This procedure wasrepeated until all the DCM solution was added. The resulting suspensionwas filtered to afford crude[4-(3-{2-[(E)-3,5-diamino-6-chloro-pyrazine-2-carbonylimino]-1,3,8-triaza-spiro[4.5]dec-8-yl}-3-oxo-propyl)-phenoxy]-aceticacid dipropylcarbamoylmethyl ester as an off-white solid. The solid wassuspended in iPrOH (50 ml) and heated to 60° C. for 3 h. A whitesuspension formed which was cooled to RT and the solid collected byfiltration, and dried under vacuum to afford the title compound; ¹H NMR(400 MHz, DMSO-d6) δ 8.42 (1, s), 8.36 (1H, s), 7.15 (2H, d), 6.86 (2H,d), 6.72 (2H, br s), 4.88 (2H, s), 4.81 (2H, s), 3.62, 3.40 (2H, m),3.56, 3.37 (2H, m), 3.38 (2H, s), 3.18-3.12 (4H, m), 2.74 (2H, t), 2.59(2H, t), 1.67-1.54 (4H, m), 1.54, 1.45 (4H, AB), 0.85 (3H, t), 0.80 (3H,t). LC-MS Rt 5.08 mins; 672.3 [M+H]⁺, Method (i)

Method B Step 1:[4-(3-{2-[(E)-3,5-Diamino-6-chloro-pyrazine-2-carbonylimino]-1,3,8-triaza-spiro[4.5]dec-8-yl}-3-oxo-propyl)-phenoxy]-aceticacid

[4-(3-{2-[(E)-3,5-Diamino-6-chloro-pyrazine-2-carbonylimino]-1,3,8-triaza-spiro[4.5]dec-8-yl}-3-oxo-propyl)-phenoxy]-aceticacid tert-butyl ester (WO09074575, Ex. 71, page 134) (28.4 g, 48.4 mmol)was slurried in 1,4-dioxane (260 ml) and stirred for 1 h. 4N HCl indioxane (121 ml, 484 mmol) was added dropwise over 15 min and thereaction mixture was stirred overnight. The resulting yellow solid wasfiltered and washed with diethyl ether. The solid was slurried in freshdiethyl ether (500 ml) and sonicated for 30 min. The resulting solid wascollected by filtration and dried in vacuo to afford the title productas a mono hydrochloride, di-dioxan solvate; LC-MS Rt 0.79 mins; 531.3and 533.3 [M+H]⁺, Method 2minLC_v003.

Step 2:[4-(3-{2-[(E)-3,5-Diamino-6-chloro-pyrazine-2-carbonylimino]-1,3,8-triaza-spiro[4.5]dec-8-yl}-3-oxo-propyl)-phenoxy]-aceticacid dipropylcarbamoylmethyl ester

A solution of[4-(3-{2-[(E)-3,5-Diamino-6-chloro-pyrazine-2-carbonylimino]-1,3,8-triaza-spiro[4.5]dec-8-yl}-3-oxo-propyl)-phenoxy]-aceticacid (Step 1) (811 mg, 1.53 mmol) in DMF (30 ml) was treated with2-chloro-N,N-dipropyl-acetamide (813 mg, 4.59 mmol) and sodium hydrogencarbonate (808 mg, 8.92 mmol). The reaction mixture was heated at 70° C.for 7 days. The reaction was allowed to cool to RT and diluted withwater (100 ml). A white precipitate formed which was extracted intoEtOAc (2×75 ml) and DCM (2×75 ml). The organic extracts were combined,dried over MgSO₄ and concentrated in vacuo to afford a brown oil.Purification by chromatography on silica eluting with a gradient of0-20% EtOH/DCM afforded a yellow oily solid which was dissolved in DCM(˜5 ml) and filtered through a 2 μM syringe filter. The solution wasconcentrated in vacuo to 1 ml, and diethyl ether (10 ml) was added. Theresulting white precipitate was collected by filtration and dried invacuo at 50° C. overnight to afford the title compound; ¹H NMR (400 MHz,DMSO-d6) δ 9.10-7.94 (1H br), 8.43 (1H br s), 8.36 (1H, br s), 7.15 (2H,d), 6.96-6.45 (1H, br s), 6.86 (2H, d), 6.72 (2H, br s), 4.88 (2H, s),4.81 (2H, s), 3.62-3.37 (4H, m), 3.38 (2H, s), 3.19-3.14 (4H, m), 2.74(2H, t), 2.59 (2H, t), 1.67-1.54 (4H, m), 1.54-1.45 (4H, m), 0.85 (3H,t), 0.80 (3H, t). LC-MS Rt 4.45 mins; 672.5 [M+H]⁺, Method 10minLC_v003.

Example 2.1[4-(3-{2-[(E)-3,5-Diamino-6-chloro-pyrazine-2-carbonylimino]-1,3,8-triaza-spiro[4.5]dec-8-yl}-3-oxo-propyl)-phenoxy]-aceticacid tert-butoxycarbonylmethyl ester

The title compound was prepared analogously to[4-(3-{2-[(E)-3,5-diamino-6-chloro-pyrazine-2-carbonylimino]-1,3,8-triaza-spiro[4.5]dec-8-yl}-3-oxo-propyl)-phenoxy]-aceticacid dipropylcarbamoylmethyl ester (Ex. 2.0) by replacing2-chloro-N,N-dipropyl-acetamide with t-butyl bromoacetate; ¹H NMR (400MHz, DMSO-d6) δ 8.45 (1H, br), 8.37 (1H, br), 7.16 (2H, d), 6.85 (2H,d), 6.8-6.6 (2H, br), 4.83 (2H, s), 4.64 (2H, s), 3.61 (1H, m), 3.58(1H, m), 3.39 (2H, s), 3.38 (1H, m), 3.30 (1H, m), 2.75 (2H, t), 2.59(2H, t), 1.60 (4H, m), 1.42 (9H, s). LC-MS Rt 4.27 mins; 645.3 [M+H]⁺,Method 10minLC_v002.

The compounds of the following tabulated Examples (Table 2) wereprepared by a similar method to that of Example 2.0 from[4-(3-{2-[(E)-3,5-Diamino-6-chloro-pyrazine-2-carbonylimino]-1,3,8-triaza-spiro[4.5]dec-8-yl}-3-oxo-propyl)-phenoxy]-aceticacid and the appropriate Intermediate (which are commerciallyavailable).

TABLE 2 Ex. Name [M + H]⁺/NMR 2.2 [4-(3-{2-[(E)-3,5-Diamino-6-chloro- 1HNMR (400 MHz, DMSO-d6) δ 8.50 pyrazine-2-carbonyl imino]-1,3,8- (2H, bs), 7.38-7.31 (5H, m), 7.14 (2H, d), triaza-spiro[4.5]dec-8-yl}-3-oxo-6.84 (2H, d), 5.19 (2H, s), 4.87 (4H, s), propyl)-phenoxy]-acetic acid3.75-3.40 (4H, m), 3.43 (2H, s), 2.75 (2H, benzyloxycarbonylmethyl estert), 2.60 (2H, t), 1.63 (4H, m). LC-MS Rt 1.29 mins; 679.4 [M + H]+ ,Method 2minLC_v002 2.3 [4-(3-{2-[(E)-3,5-Diamino-6-chloro- 1H NMR (400MHz, DMSO-d6) δ 8.50 pyrazine-2-carbonylimino]-1,3,8-triaza- (2H, b s),7.16 (2H, d), 6.87 (2H, d), 4.88 spiro[4.5]dec-8-yl}-3-oxo-propyl)- (2H,s), 4.82 (2H, s), 3.75-3.55 (2H, m), phenoxy]-acetic acid diethylcarb3.46 (2H, br s), 3.30-3.18 (4H, m, hidden amoylmethyl ester by D2Opeak), 2.76 (2H, t), 2.61 (2H, t), 1.64 (4H, br s), 1.12 (3H, t), 1.03(3H, t). LC-MS Rt 1.13 mins; 644.5 [M + H]+ , Method 2minLC_v002. 2.4[4-(3-{2-[(E)-3,5-Diamino-6-chloro- ¹H NMR (400 MHz, DMSO-d6) δ 8.63pyrazine-2-carbonylimino]-1,3,8- (2H, br s), 7.16 (2H, d), 7.20-7.00(2H, br triaza-spiro[4.5]dec-8-yl}-3-oxo- s), 6.87 (2H, d), 4.91 (2H,s), 4.81 (2H, s), propyl)-phenoxy]-acetic acid 2-oxo-2- 3.69 (1H, m),3.57 (1H, m), 3.46 (2H, s), piperidin-1-yl-ethyl ester 3.41 (2H, t),3.40-3.00 (4H, hidden by D₂O peak), 2.75 (2H, t), 2.51 (2H, t),1.63-1.43 (10H, m). LC-MS Rt 1.16 mins; 656.5 [M + H]⁺, Method2minLC_v002.

TABLE 2.2 Ex. Structure 2.2

2.3

2.4

Example 3.0[2-Chloro-4-(3-{2-[(E)-3,5-diamino-6-chloro-pyrazine-2-carbonylimino]-1,3,8-triaza-spiro[4.5]dec-8-yl}-3-oxo-propyl)-phenoxy]-aceticacid dipropylcarbamoylmethyl ester

Step 1:[2-Chloro-4-(3-{2-[(E)-3,5-diamino-6-chloro-pyrazine-2-carbonylimino]-1,3,8-triaza-spiro[4.5]dec-8-yl}-3-oxo-propyl)-phenoxy]-aceticacid tert-butyl ester

A stirred solution of3-(4-tert-butoxycarbonylmethoxy-3-chloro-phenyl)-propionic acid(Intermediate B)(4 g, 12.71 mol) in DMF (80 ml) at RT was treated withHATU (4.83 g, 12.71 mmol), N-methylmorpholine (5.73 ml, 52.1 mmol)followed by 3,5-diamino-6-chloro-pyrazine-2-carboxylic acid[1,3,8-triaza-spiro[4.5]dec-(2E)-ylidene]-amide (WO09074575, Ex. 38,page 123) (5.31 g, 13.34 mmol). The resulting mixture was stirred at RTfor 1 h and then diluted with water (500 ml). The resulting solid wascollected by filtration and rinsed with water. Purification bychromatography on silica eluting with 0-7% 7N ammonia in MeOH/DCMafforded the title product; ¹H NMR (400 MHz, DMSO-d6) δ 8.53 (2H, brhump), 7.34 (1H, d), 7.14 (1H, dd), 6.90 (1H, d), 6.87 (2H, br hump),4.74 (2H, s), 3.65 (1H, m), 3.57 (1H, m), 3.43 (2H, s), 3.37 (2H, m),2.75 (2H, t), 2.62 (2H, t), 1.62 (4H, m), 1.43 (9H, s).

LC-MS Rt 1.30 mins; 621.4 [M+H]⁺, Method 2minLC_v002.

Step 2:[2-Chloro-4-(3-{2-[(E)-3,5-diamino-6-chloro-pyrazine-2-carbonylimino]-1,3,8-triaza-spiro[4.5]dec-8-yl}-3-oxo-propyl)-phenoxy]-aceticacid

[2-Chloro-4-(3-{2-[(E)-3,5-diamino-6-chloro-pyrazine-2-carbonylimino]-1,3,8-triaza-spiro[4.5]dec-8-yl}-3-oxo-propyl)-phenoxy]-aceticacid tert-butyl ester (step 1) (5.3 g, 8.53 mmol) was suspended in1,4-dioxane (25 ml) and treated with 4N HCl in dioxane (30 ml). Thereaction mixture was stirred at RT for 24 h and filtered. The solid waswashed with iso-hexane (300 ml) to afford the title compound as ahydrochloride salt;

¹H NMR (400 MHz, DMSO-d6) δ 11.00 (1H, s), 9.39 (1H, s), 9.07 (1H, s),7.46 (2H, br s), 7.34 (1H, d), 7.14 (1H, dd), 6.92 (1H, d), 4.89 (2H,s), 3.83 (1H, m), 3.75 (1H, m), 3.70 (2H, s), 3.30 (1H, m), 3.16 (1H,m), 2.75 (2H, t), 2.64 (2H, t), 1.78 (4H, m). LC-MS Rt 1.10 mins; 565.3[M+H]⁺, Method 2minLC_v002.

Step 3:[2-Chloro-4-(3-{2-[(E)-3,5-diamino-6-chloro-pyrazine-2-carbonylimino]-1,3,8-triaza-spiro[4.5]dec-8-yl}-3-oxo-propyl)-phenoxy]-aceticacid dipropylcarbamoylmethyl ester

A solution of[2-Chloro-4-(3-{2-[(E)-3,5-diamino-6-chloro-pyrazine-2-carbonylimino]-1,3,8-triaza-spiro[4.5]dec-8-yl}-3-oxo-propyl)-phenoxy]-aceticacid (step 2)(4.82 g, 8.52 mmol) in DMF (70 ml) was treated with2-chloro-N,N-dipropyl-acetamide (3.18 g, 17.9 mmol) followed by sodiumhydrogen carbonate (2.26 g, 26.9 mmol) and the resulting suspensionheated to 60° C. overnight. Water (500 ml) was added to the reactionmixture and the product extracted with EtOAc (1000 ml). The organicextracts were washed with water (500 ml) and brine (500 ml), dried(MgSO₄) and concentrated in vacuo. Purification by chromatography onsilica eluting with 0-12% 2N ammonia in ethanol in DCM followed byfurther purification by C18 reverse phase chromatography eluting withMeCN/water afforded the title product; ¹H NMR (400 MHz, DMSO-d6) δ 8.43(1H, br s), 8.36 (1H, br s), 7.34 (1H, d), 7.14 (1H, dd), 7.02 (1H, s),6.71 (2H, br s), 4.95 (2H, s), 4.89 (2H, s), 3.67-3.53 (2H, m), 3.39(3H, br s), 3.19 (2H, t), 3.13 (2H, t), 2.75 (2H, t), 2.62 (2H, t),1.65-1.41 (8H, m), 0.85 (3H, t), 0.80 (3H, t). LC-MS Rt 4.11 mins; 706.5[M+H]⁺, Method 10minLC_v003.

Example 43-(3-{2-[(E)-3,5-Diamino-6-chloro-pyrazine-2-carbonylimino]-1,3,8-triaza-spiro[4.5]decane-8-carbonyl}-benzenesulfonylamino)-propionicacid 2-oxo-2-(2-trifluoromethyl-pyrrolidin-1-yl)-ethyl ester

Step 1:3-(3-{2-[(E)-3,5-Diamino-6-chloro-pyrazine-2-carbonylimino]-1,3,8-triaza-spiro[4.5]decane-8-carbonyl}-benzenesulfonylamino)-propionicacid tert-butyl ester

The title compound was prepared from3-(2-tert-butoxycarbonyl-ethylsulfamoyl)-benzoic acid (Int. AD) and3,5-diamino-6-chloro-pyrazine-2-carboxylic acid[1,3,8-triaza-spiro[4.5]dec-(2E)-ylidene]-amide hydrochloride in ananalogous method to Example 1; LC-MS Rt 0.93 mins; 636.3 [M+H]⁺, Method2minLC_v003.

Step 2:3-(3-{2-[(E)-3,5-Diamino-6-chloro-pyrazine-2-carbonylimino]-1,3,8-triaza-spiro[4.5]decane-8-carbonyl}-benzenesulfonylamino)-propionicacid

A solution of3-(3-{2-[(E)-3,5-diamino-6-chloro-pyrazine-2-carbonylimino]-1,3,8-triaza-spiro[4.5]decane-8-carbonyl}-benzenesulfonylamino)-propionicacid tert-butyl ester (step 1) (900 mg, 1.415 mmol) in 4N HCl in dioxane(10 ml) was stirred for 16 h at RT. Iso-hexane (10 ml) was added to thereaction mixture and resulting suspension was sonicated for 1 hour atRT. The solvent was decanted off and the solids washed with iso-hexane(10×25 ml). The resulting pale yellow crystals were dried under vacuum @45° C. for 36 h to afford the title compound; LC-MS Rt 0.80 mins; 580.0[M+H]⁺, Method 2minLCv003.

Step 3:3-(3-{2-[(E)-3,5-Diamino-6-chloro-pyrazine-2-carbonylimino]-1,3,8-triaza-spiro[4.5]decane-8-carbonyl}-benzenesulfonylamino)-propionicacid 2-oxo-2-(2-trifluoromethyl-pyrrolidin-1-yl)-ethyl ester

To a stirred solution of3-(3-{2-[(E)-3,5-Diamino-6-chloro-pyrazine-2-carbonylimino]-1,3,8-triaza-spiro[4.5]decane-8-carbonyl}-benzenesulfonylamino)-propionicacid (step 2) (400 mg, 0.690 mmol) and N,N-diisopropylethylamine (357mg, 2.76 mmol) in DMF (4 ml) was added HATU (262 mg, 0.690 mmol). Thesolution was stirred at RT for 15 min, after which time2-hydroxy-1-(2-(trifluoromethyl)pyrrolidin-1-yl)ethanone (Int. D) (262mg, 0.690 mmol) in DMF (2 ml) was added. The resulting solution wasstirred at RT for 6 days, after which time it was poured onto water (10ml). A white precipitate formed which was collected by filtration, andwashed with iso-hexane (20 ml), and purified by C18 reverse phasechromatography eluting with MeCN/water/0.1% TFA to afford the titleproduct; ¹H NMR (400 MHz, DMSO-d6) δ 11.03 (1H, s), 9.37 (1H, s), 9.02(1H, s), 7.92-7.29 (4H, m), 7.24 (¹⁴N, s) 7.12 (¹⁴N, s), 6.96 (¹⁴N, s),4.95-3.90 (11H, m), 3.66 (2H, t), 3.41 (2H, t), 3.35 (2H, m), 3.07 (2H,t) 2.20-1.61 (7H, m); LC-MS Rt 0.91 mins; 759.0 [M+H]⁺, Method2minLC_v003.

Example 4.1[2-Chloro-4-(3-{2-[(E)-3,5-diamino-6-chloro-pyrazine-2-carbonylimino]-1,3,8-triaza-spiro[4.5]dec-8-yl}-3-oxo-propyl)-phenoxy]-aceticacid 2-(2-oxo-piperidin-1-yl)-ethyl ester

The title compound was prepared by an analogous method to Example 4.0,replacing3-(3-{2-[(E)-3,5-Diamino-6-chloro-pyrazine-2-carbonylimino]-1,3,8-triaza-spiro[4.5]decane-8-carbonyl}-benzenesulfonylamino)-propionicacid with[2-Chloro-4-(3-{2-[(E)-3,5-diamino-6-chloro-pyrazine-2-carbonylimino]-1,3,8-triaza-spiro[4.5]dec-8-yl}-3-oxo-propyl)-phenoxy]-aceticacid (Ex. 3, step 2), and2-hydroxy-1-(2-(trifluoromethyl)pyrrolidin-1-yl)ethanone (Int. D) with1-(2-hydroxy-ethyl)piperidin-2-one; ¹H NMR (400 MHz, DMSO-D6) δ 11.1(1H, s), 9.34 (1H, s), 9.06 (1H, s), 7.55-6.9 (4H, m), 4.84 (4H, s),4.22 (2H, m), 3.95-3.41 (10H, m), 3.29 (2H, s), 2.77 (2H, m), 2.64 (2H,m), 2.59 (2H, m), 2.21 (2H, t), 1.88-1.59 (4H, m); LC-MS Rt 0.91 mins;690.4 [M+H]⁺, Method 2minLCv003.

Example 4.2[4-(3-{2-[(E)-3,5-Diamino-6-chloro-pyrazine-2-carbonylimino]-1,3,8-triaza-spiro[4.5]dec-8-yl}-3-oxo-propyl)-phenoxy]-aceticacid 2-morpholin-4-yl-2-oxo-ethyl ester

The title compound was prepared by an analogous method to Example 4.0,replacing3-(3-{2-[(E)-3,5-Diamino-6-chloro-pyrazine-2-carbonylimino]-1,3,8-triaza-spiro[4.5]decane-8-carbonyl}-benzenesulfonylamino)-propionicacid with[4-(3-{2-[(E)-3,5-diamino-6-chloro-pyrazine-2-carbonylimino]-1,3,8-triaza-spiro[4.5]dec-8-yl}-3-oxo-propyl)-phenoxy]-aceticacid (Intermediate B), and2-hydroxy-1-(2-(trifluoromethyl)pyrrolidin-1-yl)ethanone (Int. D) with2-hydroxy-1-morpholin-4-yl-ethanone; ¹H NMR (400 MHz, DMSO-d6) δ 8.40(2H, br hump), 7.15 (2H, d), 6.87 (2H, d), 6.72 (2H, br s), 4.93 (2H,s), 4.82 (2H, s), 3.63 (2H, m), 3.56 (4H, m), 3.42 (2H, m), 3.40 (8H,m), 2.74 (2H, t), 2.59 (2H, t), 1.59 (4H, m). LC-MS Rt 3.10 mins; 658.4[M+H]⁺, Method 10minLC_v002.

Example 5.01-[(3-{2-[(E)-3,5-Diamino-6-chloro-pyrazine-2-carbonylimino]-1,3,8-triaza-spiro[4.5]decane-8-carbonyl}-benzenesulfonylamino)-methyl]-cyclobutanecarboxylicacid dipropylcarbamoylmethyl ester

Step 1:1-[(3-{2-[(E)-3,5-Diamino-6-chloro-pyrazine-2-carbonylimino]-1,3,8-triaza-spiro[4.5]decane-8-carbonyl}-benzenesulfonylamino)-methyl]-cyclobutanecarboxylicacid tert-butyl ester

A solution of3-[(1-tert-butoxycarbonyl-cyclobutylmethyl)-sulfamoyl]-benzoic acid(Int. AC) (1.37 g, 3.71 mmol) in DMF (20 ml) was treated withN-methylmorpholine (1.631 ml, 14.83 mmol) and HATU (1.41 g, 3.71 mmol)followed by 3,5-diamino-6-chloro-pyrazine-2-carboxylic acid[1,3,8-triaza-spiro[4.5]dec-(2E)-ylidene]-amide (1.27 g, 3.89 mmol). Thereaction mixture was stirred at RT for 2 h and then partitioned betweenEtOAc and water. The organic portion was separated and washed with 0.5Maqueous 1,5,7-triazabicyclo[4.4.0]dec-5-ene solution, brine, dried overMgSO₄ and concentrated in vacuo. Purification of the resulting oil bychromatography on silica eluting with 5-8% MeOH in iso-hexane affordedthe title compound; LC-MS Rt 1.03 mins; 676.4 [M+H]⁺, Method 2minLCv003.

Step 2:1-[(3-{2-[(E)-3,5-Diamino-6-chloro-pyrazine-2-carbonylimino]-1,3,8-triaza-spiro[4.5]decane-8-carbonyl}-benzenesulfonylamino)-methyl]-cyclobutanecarboxylicacid

A suspension of1-[(3-{2-[(E)-3,5-diamino-6-chloro-pyrazine-2-carbonylimino]-1,3,8-triaza-spiro[4.5]decane-8-carbonyl}-benzenesulfonylamino)-methyl]-cyclobutanecarboxylicacid tert-butyl ester (step 1)(730 mg, 1.080 mmol) in 4N HCl in dioxane(10 ml, 40.0 mmol) was stirred at RT for 1 h. The reaction mixture waspartitioned between water and EtOAc and the pH of the aqueous portionwas adjusted to 14 by addition of 2N NaOH. The basic aqueous solutionwas washed with EtOAc and concentrated in vacuo. The dry residue wasslurried in water and the pH was adjusted to 6 by dropwise addition of2N HCl. The resulting solid was filtered and washed with water (20 ml),diethyl ether (20 ml) and dried under vacuum at 50° C. to afford thetitle product; LC-MS Rt 2.29 mins; 620.3 [M+H]⁺, Method 10minLC_v003.

Step 3:1-[(3-{2-[(E)-3,5-Diamino-6-chloro-pyrazine-2-carbonylimino]-1,3,8-triaza-spiro[4.5]decane-8-carbonyl}-benzenesulfonylamino)-methyl]-cyclobutanecarboxylicacid dipropylcarbamoylmethyl ester

A solution of1-[(3-{2-[(E)-3,5-diamino-6-chloro-pyrazine-2-carbonylimino]-1,3,8-triaza-spiro[4.5]decane-8-carbonyl}-benzenesulfonylamino)-methyl]-cyclobutanecarboxylicacid (step 2) (350 mg, 0.564 mmol) in DMF (10 ml) was stirred at 50° C.with pre-activated molecular sieves for 30 min.2-Chloro-N,N-dipropylacetamide (100 mg, 0.564 mmol), sodium bicarbonate(142 mg, 1.693 mmol) and NaI (8.46 mg, 0.056 mmol) were added andstirring continued at 50° C. After 3.5 h, the mixture was allowed tocool to RT and the molecular sieves were removed. The mixture was pouredinto water and the resulting white suspension was filtered, washed withwater and dried in vacuo. The solid was dissolved in DCM and diethylether was added to yield a white precipitate which was collected byfiltration and dried under vacuum overnight to afford the title product;¹H NMR (400 MHz, DMSO-d6) δ 9.30-7.74 (4H, br hump), 7.88 (1H, d), 7.79(1H, br s), 7.69 (1H, dd), 7.66 (1H, m), 7.11-6.67 (1H, br hump),6.84-6.61 (2H, br s), 4.76 (2H, s), 3.82-3.29 (4H, m), 3.44 (2H, s),3.18-3.14 (4H, m), 3.15 (2H, br s), 2.31-1.95 (4H, m), 1.81 (2H, m),1.83-1.65 (4H, m), 1.54-1.45 (4H, m), 0.86 (3H, t), 0.80 (3H, t); LC-MSRt 3.62 mins; 761.7 [M+H]⁺, Method 10minLC_v003.

Example 5.13-[3-(3-{2-[(E)-3,5-Diamino-6-chloro-pyrazine-2-carbonylimino]-1,3,8-triaza-spiro[4.5]decane-8-carbonyl}-phenyl)-ureido]-propionicacid dipropylcarbamoylmethyl ester

Step 1:3-[3-(3-{2-[(E)-3,5-Diamino-6-chloro-pyrazine-2-carbonylimino]-1,3,8-triaza-spiro[4.5]decane-8-carbonyl}-phenyl)-ureido]-propionicacid ethyl ester

The title compound was prepared from3-[3-(2-ethoxycarbonyl-ethyl)-ureido]-benzoic acid (Int. F) and3,5-diamino-6-chloro-pyrazine-2-carboxylic acid[1,3,8-triaza-spiro[4.5]dec-(2E)-ylidene]-amide hydrochloride using ananalogous method to Example 1; LC-MS Rt 0.81 mins; 587.4 and 589.3[M+H]⁺, Method 2minLC_v003.

Step 2:3-[3-(3-{2-[(E)-3,5-Diamino-6-chloro-pyrazine-2-carbonylimino]-1,3,8-triaza-spiro[4.5]decane-8-carbonyl}-phenyl)-ureido]-propionicacid

A solution of3-[3-(3-{2-[(E)-3,5-diamino-6-chloro-pyrazine-2-carbonylimino]-1,3,8-triaza-spiro[4.5]decane-8-carbonyl}-phenyl)-ureido]-propionicacid ethyl ester (step 1)(2.17 g, 3.70 mmol) in THF (20 ml) was treatedwith LiOH (0.167 g, 4.07 mmol) and the reaction mixture was stirred atRT for 1 h. The mixture was partitioned between EtOAc and water and theaqueous portion was acidified to neutral pH and concentrated in vacuo.The residue was suspended in water (100 ml), and the resultingsuspension was filtered, washed with cold water (100 ml) and dried toafford the title product; LC-MS Rt 0.73 mins; 559.3 [M+H]⁺, Method2minLC_v003.

Step 3:3-[3-(3-{2-[(E)-3,5-Diamino-6-chloro-pyrazine-2-carbonylimino]-1,3,8-triaza-spiro[4.5]decane-8-carbonyl}-phenyl)-ureido]-propionicacid dipropylcarbamoylmethyl ester

The title compound was prepared by an analogous method Example 5.0 byreplacing1-[(3-{2-[(E)-3,5-diamino-6-chloro-pyrazine-2-carbonylimino]-1,3,8-triaza-spiro[4.5]decane-8-carbonyl}-benzenesulfonylamino)-methyl]-cyclobutanecarboxylicacid (Ex. 4.0, step 3) with3-[3-(3-{2-[(E)-3,5-Diamino-6-chloro-pyrazine-2-carbonylimino]-1,3,8-triaza-piro[4.5]decane-8-carbonyl}-phenyl)-ureido]-propionicacid (Ex. 5.1, step 2); ¹H NMR (400 MHz, DMSO-d6) δ 8.75 (1H, s),8.50-8.40 (4H, br hump), 7.54 (1H, s), 7.30 (2H, m), 6.88 (1H, d), 6.72(2H, br s), 6.31 (1H, t), 4.78 (2H, s), 3.80-3.50 (4H, br humps), 3.43(2H, s), 3.40 (2H, br s), 3.35-3.16 (4H, m), 2.56 (2H, t), 1.80-1.60(4H, br m), 1.54-1.45 (4H, m), 0.86 (3H, t), 0.80 (3H, t); LC-MS Rt 2.85mins; 700.5 [M+H]⁺, Method 10minLCv003.

Example 5.21-[(3-{2-[(E)-3,5-Diamino-6-chloro-pyrazine-2-carbonylimino]-1,3,8-triaza-spiro[4.5]decane-8-carbonyl}-benzenesulfonylamino)-methyl]-cyclobutanecarboxylicacid 2-oxo-2-(2-trifluoromethyl-pyrrolidin-1-yl)-ethyl ester

The title compound was prepared by an analogous method to that ofExample 5.0 by replacing 2-chloro-N,N-dipropylacetamide (Ex 5.0 step 3)with 2-chloro-1-(2-(trifluoromethyl)pyrrolidin-1-yl)ethanone (Int. E);¹H NMR (400 MHz, DMSO-d6) δ 8.54 (1H, br), 8.37 (1H, br), 7.89 (1H, m),7.84 (1H, m), 7.79 (1H, m), 7.70 (1H, m), 6.73 (2H, br), 4.87 (1H, m),4.74 (2H, m), 3.82-3.36 (8H, br m), 3.70 (2H, m), 3.15 (2H, br s),2.10-1.63 (12H, br m); LC-MS Rt 3.27 mins; 799.4 [M+H]⁺, Method10minLC_v003.

Preparation of Intermediate Compounds Intermediate AA3-(2-Dipropylcarbamoylmethoxycarbonyl-ethylsulfamoyl)-benzoic acid

The title compound may be prepared by either Method A or B:

Method A Step 1: 3-Benzyloxycarbonylamino-propionic aciddipropylcarbamoylmethyl ester

To a solution of benzyloxycarbonylamino-propionic acid (22.3 g, 99.9mmol) in DMF (150 ml) was added potassium carbonate (19.3 g, 139.9mmol). 2-Chloro-N,N-dipropyl-acetamide (17.7 g, 99.9 mmol) was addedover 30 min, and the reaction mixture was heated to 60° C. and stirredfor 2.5 h. The reaction mixture was allowed to cool to RT and dilutedwith water (500 ml) and extracted with isopropyl acetate (total 500 ml).The combined organic phases were washed with water (3×200 ml) to afforda solution of 3-benzyloxycarbonylamino-propionic aciddipropylcarbamoylmethyl ester in isopropyl acetate which was notisolated further. LC-MS; 365.2 [M+H]⁺ Method (i)

Step 2: 3-Amino-propionic acid dipropylcarbamoylmethyl estertrifluoroacetate

A solution of 3-benzyloxycarbonylamino-propionic aciddipropylcarbamoylmethyl ester in isopropyl acetate (33.2 g, 91.0 mmol in129.9 g total mass of solution) was treated with TFA (7.05 ml, 92.0mmol) while the internal temperature was maintained at 20° C., followedby 10% Pd/C (3.3 g, 50% wet) and stirred under an atmosphere of H₂ (3atm) for 3.5 h to afford 3-amino-propionic acid dipropylcarbamoylmethylester trifluoroacetate. The solution was used directly in the nextreaction without isolation.

Step 3: Benzyl3-(2-Dipropylcarbamoylmethoxycarbonyl-ethylsulfamoyl)-benzoic acidbenzyl ester

A solution of 3-amino-propionic acid dipropylcarbamoylmethyl estertrifluoroacetate (25.4 g 73.8 mmol) in isopropyl acetate was cooled to0° C. and treated with N-methylmorpholine (26.3 g, 221.5 mmol), water(40 ml) and DMAP (90.4 mg, 0.74 mmol). 3-Chlorosulfonyl-benzoic acidbenzyl ester (24.1 g, 77.5 mmol) in isopropyl acetate (44 ml) was addedand the reaction mixture was stirred for 2 h at 0-5° C. The layers wereseparated and the organic phase was washed with sat. aq. NaHCO₃ (3×26ml), diluted with water (10 ml), the pH adjusted to 6 with 1N HClsolution, and washed with brine. The resulting solution was concentratedto provide a solution of benzyl3-(2-Dipropylcarbamoylmethoxycarbonyl-ethylsulfamoyl)-benzoic acidbenzyl ester in isopropyl acetate which was used in further reactionswithout isolation. LC-MS; [M+H]⁺505.1 Method (i)

Step 4: Synthesis of3-(2-Dipropylcarbamoylmethoxycarbonyl-ethylsulfamoyl)-benzoic acid

A solution of benzyl3-(2-Dipropylcarbamoylmethoxycarbonyl-ethylsulfamoyl)-benzoic acidbenzyl ester (5 g, 33.25 mmol) in isopropyl acetate (82 ml) was treatedwith 10% Pd/C (0.84 g, 50% wet), and stirred under H₂ (3 atm) overnight.The catalyst was removed by filtration. To the filtrate was added 10%Pd/C (1.68 g, 50% wet), and the reaction stirred under H₂ (3 atm) for 18h. The catalyst was removed by filtration and further 10% Pd/C (1.68 g,50% wet) was added and the reaction stirred under H₂ (1 atm) for 18 h.The catalyst was removed by filtration and washed with isopropyl acetate(20 ml). The combined filtrates were concentrated in vacuo and heptanewas added to the solution and stirred at RT for 2 hr then −2° C. for 4h. The solid which formed was collected by filtration and dried undervacuum at 40° C. to afford the title compound; LC-MS; 415.1 [M+H]⁺,Method (i).

Method B Step 1: 3-tert-Butoxycarbonylamino-propionic aciddipropylcarbamoylmethyl ester

To a stirred suspension of Boc-Beta-Ala-OH (40.0 g, 211 mmol) in DMF(200 ml) at 60° C. under N₂ was added potassium carbonate (40.0 g, 289mmol). To this mixture was added 2-chloro-N,N-dipropyl-acetamide (36.7g, 207 mmol) in DMF (75 ml). The reaction mixture was allowed to stir at60° C. overnight. The reaction was allowed to cool to RT and dilutedwith DCM (400 ml) followed by water (500 ml). The organic layer wasseparated and washed with brine (200 ml), dried over MgSO₄ andconcentrated in vacuo to yield a pale yellow oil. To the oil was addedn-heptane (500 ml) (azeotrope for DMF) which was concentrated in vacuoto afford the title compound; LC-MS Rt 1.14 mins; 331.3 [M+H]⁺, Method2minLC_v003.

Step 2: 3-Amino-propionic acid dipropylcarbamoylmethyl ester

To a cooled stirred solution of 3-tert-butoxycarbonylamino-propionicacid dipropylcarbamoylmethyl ester (step 1)(36.5 g, 110 mmol) in drydioxane under N₂ was added dropwise 4N HCl in dioxane (18.12 ml, 597mmol). The resulting mixture was allowed to warm to RT and stirredovernight. The solvent was removed in vacuo and the crude product wassuspended in EtOAc (500 ml) and sonicated for 1 h. The resulting whiteprecipitate was isolated by filtration and was dried under vacuum at 40°C. for 1 h to afford a pale yellow solid. Recrystallisation from EtOAcafforded the title compound; LC-MS Rt 0.77 mins; 231.2 [M+H]⁺, Method2minLC_v003.

Step 3: 3-(2-Dipropylcarbamoylmethoxycarbonyl-ethylsulfamoyl)-benzoicacid

To a stirred solution of 3-amino-propionic acid dipropylcarbamoylmethylester (step 2) (20.1 g, 75 mmol) in DCM (240 ml) at 0° C. under N₂ wasadded DMAP (0.46 g, 3.76 mmol) followed by TEA (38.8 ml, 278 mmol). Thereaction mixture was treated with a solution of 3-chlorosulfonylbenzoicacid (16.6 g, 75 mmol) in DCM (200 ml). The mixture was allowed to stirat 0° C. for 1 h and then warmed up to RT for 1 h. Water (200 ml) wasadded and the pH adjusted with 1N HCl (100 ml). The organic layer wasseparated, dried over MgSO₄ and concentrated in vacuo. Purification byC18 reverse phase chromatography, eluting with MeCN/water (1% HCl)afforded the title product; LC-MS Rt 1.01 mins; 415.2 [M+H]⁺, Method2minLC_v003.

Intermediate AB3-(2-Cyclohexyloxycarbonylmethoxycarbonyl-ethylsulfamoyl)-benzoic acid

Step 1: 3-tert-Butoxycarbonylamino-propionic acidcyclohexyloxycarbonylmethyl ester

This compound was prepared from cyclohexyl 2-chloroacetate andBoc-Beta-Ala-OH by an analogous method to3-tert-butoxycarbonylamino-propionic acid dipropylcarbamoylmethyl ester(Int. AA step 1). Cesium carbonate was used in place of potassiumcarbonate; ¹H NMR (400 MHz, DMSO-d6) δ 6.83 (1H, br s), 4.76-4.70 (1H,m), 4.62 (2H, s), 3.20-3.12 (2H, m), 2.53 (2H, under DMSO peak),1.28-1.72 (2H, m), 1.68-1.60 (2H, m), 1.53-1.18 (15H, m).

Step 2: 3-Amino-propionic acid cyclohexyloxycarbonylmethyl ester

The title compound was prepared from3-tert-butoxycarbonylamino-propionic acid cyclohexyloxycarbonylmethylester (step 1) analogously to 3-amino-propionic aciddipropylcarbamoylmethyl ester (Int. AA, step 2); ¹H NMR (400 MHz,DMSO-d6) δ 7.86 (3H, br s), 4.76-4.71 (1H, m), 4.69 (2H, s), 3.04 (2H,m), 2.77 (2H, m), 1.82-1.75 (2H, m), 1.70-1.60 (2H, m), 1.53-1.19 (6H,m).

Step 3:3-(2-Cyclohexyloxycarbonylmethoxycarbonyl-ethylsulfamoyl)-benzoic acid

A stirred solution of 3-amino-propionic acid cyclohexyloxycarbonylmethylester (step 2) (2.30 g, 10.03 mmol) in DCM (200 ml) at RT was treatedwith 3-chlorosulfonyl-benzoic acid (3.64 g, 16.50 mmol) followed byN,N-diisopropylethylamine (3.04 ml, 17.45 mmol). The resulting mixturewas stirred at RT for 1 h. The reaction mixture was washed with 2N HCl(200 ml), dried over MgSO₄ and concentrated in vacuo to afford the titlecompound; LC-MS Rt 1.34 mins; 414.2 [M+H]⁺, Method 2minLC_v002.

Intermediate AC3-[(1-tert-Butoxycarbonyl-cyclobutylmethyl)-sulfamoyl]-benzoic acid

Step 1: 1-Cyano-cyclobutanecarboxylic acid tert-butyl ester

A solution of tert-butyl 2-cyanoacetate (4.0 g, 28.3 mmol),1,3-dibromopropane (6.29 g, 31.2 mmol) in methyl ethyl ketone (100 ml)was treated with potassium carbonate (11.75 g, 85 mmol). The reactionmixture was heated to 80° C. for 17 h and then NaI (catalyst) (0.21 g,1.42 mmol) was added. The mixture was heated at 80° C. for 6 days andthen allowed to cool to RT. The mixture was filtered through Celite®(filter material) and the filtrate was concentrated in vacuo.Purification by chromatography on silica eluting with 10% EtOAc iniso-hexane afforded the title product as a colourless oil; ¹H NMR (400MHz, CDCl₃) δ 2.64 (4H, m), 2.22 (2H, m), 1.53 (9H, s).

Step 2: 1-Aminomethyl-cyclobutanecarboxylic acid tert-butyl ester

A suspension of Raney Ni (0.6 g, 13.19 mmol) in MeOH (30 ml) in ahydrogenation steel pressure vessel was treated with1-cyano-cyclobutanecarboxylic acid tert-butyl ester (2.39 g, 13.19mmol). The reaction mixture was placed under an atmosphere of hydrogen(3 bar) overnight. The mixture was filtered and concentrated in vacuo.The residue was dissolved in MeOH and passed through a 20 g Isolute®SCX-2 cartridge. The cartridge was washed with MeOH followed by product7M ammonia in MeOH. The ammonia washings were concentrated in vacuo toafford the title product which was used without further purification. ¹HNMR (400 MHz, CDCl₃) δ 2.45 (2H, m), 2.01-1.67 (6H, m), 0.49 (9H, s).

Step 3: 3-[(1-tert-Butoxycarbonyl-cyclobutylmethyl)-sulfamoyl]-benzoicacid methyl ester

A solution of 1-aminomethyl-cyclobutanecarboxylic acid tert-butyl ester(step 2) (1.47 g, 7.38 mmol) in DCM (40 ml) was treated with TEA (2.57ml, 18.44 mmol) followed by methyl 3-(chlorosulfonyl)benzoate (1.91 g,8.11 mmol) and DMAP (0.09 g, 0.74 mmol). The mixture was stirred at RTfor 2 h and then partitioned between DCM and water. The aqueous portionwas extracted with DCM and the combined organic extracts were washedwith 0.1N HCl, brine, dried over MgSO₄ and concentrated in vacuo.Purification of the resulting oil by chromatography on silica elutingwith 0-40% EtOAc in iso-hexane afforded the title compound as acolourless oil; ¹H NMR (400 MHz, CDCl₃) δ 8.56 (1H, s), 8.27 (1H, d),8.08 (1H, d), 7.64 (1H, t), 5.21 (1H, br t), 3.98 (3H, s), 3.20 (2H, d),2.37 (2H, m), 1.96 (4H, m), 1.46 (9H, s).

Step 4: 3-[(1-tert-Butoxycarbonyl-cyclobutylmethyl)-sulfamoyl]-benzoicacid

A solution of3-[(1-tert-butoxycarbonyl-cyclobutylmethyl)-sulfamoyl]-benzoic acidmethyl ester (step 3) (1.87 g, 4.88 mmol) in THF (25 ml) was treatedwith 2M NaOH (aq) (24.38 ml, 48.8 mmol) and stirred at RT for 2 h. Thereaction mixture was partitioned between diethyl ether and water. Theaqueous portion was separated and acidified to pH 1 with 2N HCl, andextracted with EtOAc, and the combined organic portions were washed withbrine, dried over MgSO₄ and concentrated in vacuo to afford the titleproduct; ¹H NMR (400 MHz, CDCl₃) δ 8.64 (1H, s), 8.33 (1H, d), 8.12 (1H,d), 7.67 (1H, t), 5.40 (1H, t), 3.24 (2H, d), 2.38 (2H, m), 1.97 (4H,m), 1.48 (9H, s). LC-MS Rt 1.13 mins; 314.1 [M+H]⁺ (-tBu), Method2minLC_v003.

Intermediate AD 3-(2-tert-Butoxycarbonyl-ethylsulfamoyl)-benzoic acid

Step 1: 3-(2-tert-Butoxycarbonyl-ethylsulfamoyl)-benzoic acid methylester

To a stirred solution of methyl 3-(chlorosulfonyl)benzoate (5.0 g, 21.31mmol) in pyridine (50 ml), was added dropwise a solution of tert-butyl3-aminopropanoate hydrochloride (5.11 g, 23.44 mmol) in pyridine (10 ml)at 0° C. The resulting solution was allowed to warm to RT and stirredfor 4 h. 1N HCl (50 ml) was added to the reaction mixture and theaqueous phase was separated and extracted with DCM (3×20 ml). Thecombined organic fractions were dried (MgSO₄) and concentrated in vacuoto afford a viscous amber oil. Purification by chromatography on silicaeluting with 0-100% EtOAc in iso-hexane afforded the title compound as acolourless oil. ¹H NMR (400 MHz, CDCl₃) δ 8.53 (1H, s), 8.27 (1H, d),8.08 (1H, d), 7.63 (1H, t) 5.38 (1H, t), 3.97 (3H, s), 3.20 (2H, m),2.48 (2H, t), 1.43 (9H, s).

Step 2: 3-(2-tert-Butoxycarbonyl-ethylsulfamoyl)-benzoic acid

A solution of 3-(2-tert-Butoxycarbonyl-ethylsulfamoyl)-benzoic acidmethyl ester (step 1)(5.0 g, 14.56 mmol) and lithium iodide (2.53 g,18.93 mmol) in pyridine (60 ml) was heated at reflux for 16 h. Thereaction mixture was concentrated in vacuo, diluted with DCM (60 ml) andpoured into 1.5N HCl (60 ml). The aqueous phase was separated andextracted with DCM (6×60 ml). The combined organic extracts were dried(MgSO₄) and concentrated in vacuo. The crude residue was purified usingan Isolute® PE-AX 10 g cartridge (anion exchange column), eluting withMeOH (200 ml) and then a 1:1 mixture of MeOH/AcOH (200 ml) to afford thetitle compound as white crystals; ¹H NMR (400 MHz, CDCl₃) δ 8.63 (1H,s), 8.32 (1H, d), 8.14 (1H, d), 7.68 (1H, t), 5.63 (1H, t), 3.22 (2H,m), 2.51 (2H, t), 1.43 (9H, s).

Intermediate B3-(4-tert-Butoxycarbonylmethoxy-3-chloro-phenyl)-propionic acid

Step 1: 3-(3-Chloro-4-hydroxy-phenyl)-propionic acid benzyl ester

A solution of 3-(4-hydroxy-phenyl)-propionic acid benzyl ester (13.36 g,52.1 mmol) in MeCN (100 ml) was cooled to 0° C. Trichloroisocyanuricacid (4.00 g, 17.2 mmol) was added, and the resulting white suspensionwas gradually allowed to warm to RT and stirred overnight. The reactionmixture was diluted with water (200 ml) and the product was extractedinto EtOAc (250 ml) and dried (MgSO₄). Purification by chromatography onsilica eluting with 0-20% EtOAc in iso-hexane afforded the titleproduct; ¹H NMR (400 MHz, DMSO-d6) δ 9.90 (1H, s), 7.32 (3H, m), 7.29(2H, d), 7.18 (1H, d), 6.97 (1H, dd), 6.85 (1H, d), 5.07 (2H, s), 2.76(2H, t), 2.64 (2H, t). LC-MS Rt 1.46 mins; 313.2 [M+H]⁺, Method2minLC_v002.

Step 2: 3-(4-tert-Butoxycarbonylmethoxy-3-chloro-phenyl)-propionic acidbenzyl ester

3-(3-Chloro-4-hydroxy-phenyl)-propionic acid benzyl ester (step 1) (5.9g, 20.3 mmol) in DMF (60 ml) was treated with potassium carbonate (5.61g, 40.6 mmol) followed by t-butyl bromoacetate (4.49 ml, 30.4 mmol) andthe mixture was heated at 60° C. overnight. The reaction mixture wasdiluted with water (500 ml), and the product extracted into EtOAc (450ml). The organic portion was washed with water (200 ml), brine (200 ml),dried over MgSO₄ and the solvent removed in vacuo to yield a yellow oil.Purification by chromatography on silica eluting with 0-20% EtOAc iniso-hexane afforded the title product; ¹H NMR (400 MHz, DMSO-d6) δ 7.34(6H, m), 7.11 (1H, d), 6.89 (1H, d), 5.07 (2H, s), 4.74 (2H, s), 2.81(2H, t), 2.68 (2H, t), 1.42 (9H, s). LC-MS Rt 1.64 mins; 427.3 [M+Na]⁺,Method 2minLC_v002.

Step 3: 3-(4-tert-Butoxycarbonylmethoxy-3-chloro-phenyl)-propionic acid

A solution of 3-(4-tert-Butoxycarbonylmethoxy-3-chloro-phenyl)-propionicacid benzyl ester (step 2) (6.35 g, 15.7 mmol) in THF (60 ml) was addedto a suspension of palladium on activated carbon (10% wt, 0.84 g, 0.78mmol) in THF (35 ml) under an inert atmosphere. The resultant mixturewas placed under an atmosphere of hydrogen (0.35 bar) for 3 h. Thereaction mixture was filtered through Celite® (filter material) andwashed through with THF (100 ml). The filtrate was concentrated in vacuoto afford the title compound as a yellow oil; ¹H NMR (400 MHz, DMSO-d6)δ 12.15 (1H, s), 7.31 (1H, d), 7.12 (1H, dd), 6.91 (1H, d), 4.74 (2H,s), 2.75 (2H, t), 2.51 (2H, t), 1.38 (9H, s).

Intermediate CA3-(4-{[(2-Hydroxy-ethyl)-methyl-carbamoyl]-methoxycarbonylmethoxy}-phenyl)-propionicacid

Step 1: 3-(4-tert-Butoxycarbonylmethoxy-phenyl)-propionic acid benzylester

This compound was prepared analogously to3-(4-tert-butoxycarbonylmethoxy-3-chloro-phenyl)-propionic acid benzylester (Int. B step 2) by replacing3-(3-chloro-4-hydroxy-phenyl)-propionic acid benzyl ester with3-(4-hydroxy-phenyl)-propionic acid benzyl ester (commerciallyavailable).

Step 2: 3-(4-Carboxymethoxy-phenyl)-propionic acid benzyl ester

3-(4-tert-Butoxycarbonylmethoxy-phenyl)-propionic acid benzyl ester(step 1) (10 g, 27 mmol) in DCM (100 ml) was treated with TFA (20 ml)and heated at reflux for 4 h. The resulting mixture was concentrated invacuo. The residue was dissolved in DCM and concentrated in vacuo afurther four times until a viscous oil of the title compound wasobtained.

Step 3:3-(4-{[(2-Hydroxy-ethyl)-methyl-carbamoyl]-methoxycarbonylmethoxy}-phenyl)-propionicacid benzyl ester

A mixture comprising 3-(4-carboxymethoxy-phenyl)-propionic acid benzylester (Step 2) (0.65 g, 1.93 mmol) and2-chloro-N-(2-hydroxy-ethyl)-N-methyl-acetamide (0.35 g, 2.31 mmol) inDMF (10 ml) and sodium hydrogen carbonate (2.5 equivalents) was stirredat RT for 2 days and concentrated in vacuo. The crude residue waspurified by chromatography on silica eluting with 0-5% MeOH in DCM toafford the title product as a viscous oil.

Step 4:3-(4-{[(2-Hydroxy-ethyl)-methyl-carbamoyl]-methoxycarbonylmethoxy}-phenyl)-propionicacid

A solution of3-(4-{[(2-hydroxy-ethyl)-methyl-carbamoyl]-methoxycarbonylmethoxy}-phenyl)-propionicacid benzyl ester (step 3) (0.44 g, 1.02 mmol) in MeOH (20 ml) wastreated with palladium on activated carbon (10% wt, 50 mg, 0.047 mmol)and stirred under an atmosphere of hydrogen for 20 h at RT. The reactionmixture was filtered through Celite® (filter material) and washedthrough with EtOAc. The filtrate was concentrated in vacuo andpurification of the crude product by chromatography on silica elutingwith 0-10% MeOH in DCM afforded the title compound; LC-MS: 340.2 [M+H]⁺

Intermediate CB3-(4-Cyclohexyloxycarbonylmethoxycarbonylmethoxy-phenyl)-propionic acid

This compound was prepared analogously to Intermediate CA by replacing2-chloro-N-(2-hydroxy-ethyl)-N-methyl-acetamide (step 3) withchloro-acetic acid cyclohexyl ester; LC-MS: 363 [M+H]⁺.

Intermediate CC3-(4-Dimethylcarbamoylmethoxycarbonylmethoxy-phenyl)-propionic acid LE

This compound was prepared analogously to Intermediate CA by replacing2-chloro-N-(2-hydroxy-ethyl)-N-methyl-acetamide (step 3) with2-chloro-N,N-dimethyl acetamide; LC-MS: 400.2 [M+H]⁺.

Intermediate D 2-Hydroxy-1-(2-(trifluoromethyl)pyrrolidin-1-yl)ethanone

A solution of 2,2-dimethyl-1,3-dioxolan-4-one (0.57 g, 5.05 mmol) and(+/−)-2-(trifluoromethyl)pyrrolidine (1.0 g, 7.19 mmol) in PhMe (20 ml)was heated at reflux for 16 h.

The reaction mixture was poured into 1N HCl (20 ml) and DCM (20 ml). Theaqueous phase was separated and extracted using DCM (3×20 ml) and thecombined organic fractions dried (MgSO₄) and concentrated in vacuo toafford the title compound. ¹H NMR (400 MHz, CDCl₃) δ 4.83 (1H, m), 4.20(2H, s), 3.5 (1H, m), 3.40 (1H, m), 3.22 (1H, s), 2.21 (2H, m), 2.10(2H, m).

Intermediate E 2-Chloro-1-(2-(trifluoromethyl)pyrrolidin-1-yl)ethanone

A solution of 2-(trifluoromethyl)pyrrolidine (500 mg, 3.59 mmol) in DCM(10 ml) was treated with chloroacetyl chloride (0.31 ml, 3.77 mmol) andstirred at RT for 1 h. The resulting mixture was concentrated in vacuoto afford the title compound as a yellow oil which was used withoutfurther purification; ¹H NMR (400 MHz, CDCl₃) δ 4.84 (1H, m), 4.11 (2H,s), 3.71 (2H, m), 2.23 (2H, m), 2.07 (2H, m).

Intermediate F 3-[3-(2-Ethoxycarbonyl-ethyl)-ureido]-benzoic acid

A solution of 3-amino benzoic acid (5.0 g, 36.5 mmol) in THF (250ml)/DCM (250 ml) was cooled to 0° C. and treated with triphosgene (3.79g, 12.76 mmol) followed by TEA (12.70 ml, 91 mmol). After 5 min, asuspension of beta-alanine ethyl ester hydrochloride (11.20 g, 72.9mmol) and TEA (12.70 ml, 91.00 mmol) in THF (10 ml)/DCM (10 ml) wereadded and the resulting mixture was allowed to warm to RT and stirredovernight. The reaction mixture was diluted with DCM (100 ml) and the pHwas adjusted to 8 by addition of sat. aq. NaHCO₃. The mixture wasextracted with water (2×250 ml). The combined aqueous extracts wereacidified to pH 1 by addition of 2N HCl, extracted with DCM and thecombined organic portions were washed with brine, dried over MgSO₄ andconcentrated in vacuo to afford the title product as an off-white solid;LC-MS Rt 0.84 mins; 281.2 [M+H]⁺, Method 2minLC_v003.

From the foregoing it will be appreciated that, although specificembodiments of the invention have been described herein for purposes ofillustration, various modifications may be made without deviating fromthe spirit and scope of the invention. Accordingly, the invention is notlimited except as by the appended claims.

EMBODIMENT/CONSISTORY CLAUSES Embodiment 1

A compound of Formula I:

or a pharmaceutically acceptable salt or solvate thereof, whereinR¹ is H, halogen, C₁-C₈-alkyl, C₁-C₈-haloalkyl, C₁-C₈-haloalkoxy,C₃-C₁₅-carbocyclic group, nitro, cyano, a C₆-C₁₅-membered aromaticcarbocyclic group, or a C₁-C₈-alkyl substituted by a C₆-C₁₅-memberedaromatic carbocyclic group;R², R³, R⁴ and R⁵ are each independently selected from H and C₁-C₆alkyl;R⁶, R⁷, R⁸ and R⁹ are each independently selected from H; SO₂R¹⁶; aryloptionally substituted by one or more Z groups; a C₃-C₁₀ carbocyclicgroup optionally substituted by one or more Z groups; C₃-C₁₄heterocyclic group optionally substituted by one or more Z groups; C₁-C₈alkyl optionally substituted by an aryl group which is optionallysubstituted by one or more Z groups, a C₃-C₁₀ carbocyclic groupoptionally substituted by one or more Z groups or a C₃-C₁₄ heterocyclicgroup optionally substituted by one or more Z groups;R¹⁰ is represented by the formula 2:

—(C₀-C₃alkylene)-B—X—(CR^(11a)R^(12a))_(m)—(CR^(11b)R^(12b))_(n)—(CR^(11c)R^(12c))_(p)-C(O)OR¹³,

wherein the alkylene groups are optionally substituted by one or more Zgroups;B is aryl optionally substituted by one or more Z groups;X is selected from a bond, —NR¹⁵(SO₂)—, —(SO₂)NR¹⁵—, —(SO₂)—,—NR¹⁵C(O)—, —C(O)NR¹⁵—, —NR¹⁵C(O)NR¹⁷—, —NR¹⁵C(O)O—, —NR¹⁵—, C(O)O,OC(O), C(O), O and S;R^(11a), R^(11b), R^(11c), R^(12a), R^(12b) and R^(12c) are eachindependently selected from H and C₁-C₆ alkyl; orR^(11a) and R^(12a) together with the carbon atom to which they areattached form a 3- to 8-membered cycloalkyl group; orR^(11b) and R^(12b) together with the carbon atom to which they areattached form a 3- to 8-membered cycloalkyl group; orR^(11c) and R^(12c) together with the carbon atom to which they areattached form a 3- to 8-membered cycloalkyl group;R¹³ is selected from (C₁-C₃ alkyl)-C(O)NR²²R²³; (C₁-C₃ alkyl)-C(O)OR²³;and (C₁-C₃ alkyl)-NR²³C(O)R²²R¹⁵ and R¹⁷ are each independently selected from H and C₁-C₆ alkyl;R¹⁶ is selected from C₁-C₈ alkyl, aryl and a 3- to 14-memberedheterocyclic group, the heterocyclic group including one or moreheteroatoms selected from N, O and S;Z is independently selected from —OH, aryl, —O-aryl, C₇-C₁₄ aralkyl,—O—C₇-C₁₄ aralkyl, C₁-C₆ alkyl, C₁-C₆ alkoxy, —NR¹⁹(SO₂)R²¹,—(SO₂)NR¹⁹R²¹, —(SO₂)R²⁰, —NR¹⁹C(O)R²⁰, —C(O)NR¹⁹R²⁰, —NR¹⁹C(O)NR²⁰R¹⁸,—NR¹⁹C(O)OR²⁰, —NR¹⁹R²¹, C(O)OR¹⁹, —C(O)R¹⁹, SR¹⁹, —OR¹⁹, oxo, CN, NO₂,and halogen, wherein the alkyl, alkoxy, aralkyl and aryl groups are eachoptionally substituted by one or more substituents selected from OH,halogen, C₁-C₄ haloalkyl and C₁-C₄ alkoxy;R¹⁸, R²⁰ and R²² are each independently selected from H and C₁-C₆ alkyl;R¹⁹, R²¹ and R²³ are each independently selected from H; C₁-C₈ alkyl;C₃-C₈ cycloalkyl; C₁-C₄ alkoxy-C₁-C₄ alkyl; (C₀-C₄ alkyl)-aryloptionally substituted by one or more groups selected from C₁-C₆ alkyl,C₁-C₆ alkoxy and halogen; (C₀-C₄ alkyl)-3- to 14-membered heterocyclicgroup, the heterocyclic group including one or more heteroatoms selectedfrom N, O and S, optionally substituted by one or more groups selectedfrom halogen, oxo, C₁-C₆ alkyl and —C(O)C₁-C₆ alkyl; —(C₀-C₄alkyl)-O-aryl optionally substituted by one or more groups selected fromC₁-C₆ alkyl, C₁-C₆ alkoxy and halogen; and (C₀-C₄ alkyl)-O-3- to14-membered heterocyclic group, the heterocyclic group including one ormore heteroatoms selected from N, O and S, optionally substituted by oneor more groups selected from halogen, C₁-C₆ alkyl and —C(O)C₁-C₆ alkyl;wherein the alkyl groups are optionally substituted by one or morehalogen atoms, hydroxyl, C₁-C₄ alkoxy, —C(O)NH₂, —C(O)NHC₁-C₆ alkyl or—C(O)N(C₁-C₆ alkyl)₂; orR¹⁹ and R²⁰ together with the nitrogen atom to which they are attachedform a 5- to 10-membered heterocyclic group, the heterocyclic groupincluding one or more further heteroatoms selected from N, O and S, theheterocyclic group being optionally substituted by one or moresubstituents selected from OH; halogen; aryl; 5- to 10-memberedheterocyclic group including one or more heteroatoms selected from N, Oand S; —S(O)₂-aryl; —S(O)₂—C₁-C₆ alkyl; C₁-C₆ alkyl optionallysubstituted by one or more halogen atoms; C₁-C₆ alkoxy optionallysubstituted by one or more OH groups or C₁-C₄ alkoxy; and C(O)OC₁-C₆alkyl, wherein the aryl and heterocyclic substituent groups arethemselves optionally substituted by C₁-C₆ alkyl, C₁-C₆ haloalkyl orC₁-C₆ alkoxy; orR¹⁹ and R²¹ together with the nitrogen atom to which they are attachedform a 5- to 10-membered heterocyclic group, the heterocyclic groupincluding one or more further heteroatoms selected from N, O and S, theheterocyclic group being optionally substituted by one or moresubstituents selected from OH; halogen; aryl; 5- to 10-memberedheterocyclic group including one or more heteroatoms selected from N, Oand S; —S(O)₂-aryl; —S(O)₂—C₁-C₆ alkyl; C₁-C₆ alkyl optionallysubstituted by one or more halogen atoms; C₁-C₆ alkoxy optionallysubstituted by one or more OH groups or C₁-C₄ alkoxy; and —C(O)OC₁-C₆alkyl, wherein the aryl and heterocyclic substituent groups arethemselves optionally substituted by C₁-C₆ alkyl, C₁-C₆ haloalkyl orC₁-C₆ alkoxy; orR¹⁸ and R²⁹ together with the nitrogen atom to which they are attachedform a 5- to 10-membered heterocyclic group, the heterocyclic groupincluding one or more further heteroatoms selected from N, O and S, theheterocyclic group being optionally substituted by one or moresubstituents selected from OH; halogen; aryl; 5- to 10-memberedheterocyclic group including one or more heteroatoms selected from N, Oand S; —S(O)₂-aryl; —S(O)₂—C₁-C₆ alkyl; C₁-C₆ alkyl optionallysubstituted by one or more halogen atoms; C₁-C₆ alkoxy optionallysubstituted by one or more OH groups or C₁-C₄ alkoxy; and —C(O)OC₁-C₆alkyl, wherein the aryl and heterocyclic substituent groups arethemselves optionally substituted by C₁-C₆ alkyl, C₁-C₆ haloalkyl orC₁-C₆ alkoxy; orR²² and R²³ together with the atom(s) to which they are attached form a5- to 10-membered heterocyclic group, the heterocyclic group includingone or more further heteroatoms selected from N, O and S, theheterocyclic group being optionally substituted by one or more Z groups;m is 0, 1, 2 or 3;n is 0, 1, 2 or 3;p is 0, 1, 2 or 3;wherein at least one of m, n or p is not 0.

Embodiment 2

A compound according to embodiment 1, wherein R¹ is halogen.

Embodiment 3

A compound according to embodiment 1 or 2, wherein R², R³, R⁴ and R⁵ areH.

Embodiment 4

A compound according to any preceding claim wherein R⁶, R⁷, R⁸ and R⁹are H.

Embodiment 5

A compound according to any preceding claim wherein B is phenyloptionally substituted by halogen.

Embodiment 6

A compound according to any preceding claim wherein X is selected from—(SO₂)NR¹⁵—, —NR¹⁵C(O)NR¹⁷—, and O.

Embodiment 7

A compound according to any preceding claim wherein R^(11a), R^(11b),R^(11c), R^(12a), R^(12b) and R^(12c) are each independently selectedfrom H and C₁-C₃ alkyl; or

R^(11a) and R^(12a) together with the carbon atom to which they areattached form a 3, 4- or 5-membered cycloalkyl group; orR^(11b) and R^(12b) together with the carbon atom to which they areattached form a 3, 4- or 5-membered cycloalkyl group; orR^(11c) and R^(12c) together with the carbon atom to which they areattached form a 3, 4- or 5-membered cycloalkyl group.

Embodiment 8

A compound according to any preceding claim wherein the sum of m, n andp is 1, 2, or, 3.

Embodiment 9

A compound according to any preceding claim wherein R¹³ is selected from(C₁ alkyl)-C(O)NR²²R²³; (C₁ alkyl)-C(O)OR²³; and (C₂ alkyl)-NR²³C(O)R²²

Embodiment 10

A compound according to any preceding claim wherein R¹³ is selected from(C₁-C₃ alkyl)-C(O)NR²²R²³ and (C₁-C₃ alkyl)-C(O)OR²³; and wherein

R²² is selected from H and C₁-C₃ alkyl;R²³ is selected from H; C₁-C₈ alkyl; C₃-C₈ cycloalkyl; C₁-C₄alkoxy-C₁-C₄ alkyl; (C₀-C₄ alkyl)-aryl optionally substituted by one ormore groups selected from C₁-C₆ alkyl, C₁-C₆ alkoxy and halogen; (C₀-C₄alkyl)-3- to 14-membered heterocyclic group, the heterocyclic groupincluding one or more heteroatoms selected from N, O and S, optionallysubstituted by one or more groups selected from halogen, oxo, C₁-C₆alkyl and C(O)C₁-C₆ alkyl; (C₀-C₄ alkyl)-O-aryl optionally substitutedby one or more groups selected from C₁-C₆ alkyl, C₁-C₆ alkoxy andhalogen; and (C₀-C₄ alkyl)-O-3- to 14-membered heterocyclic group, theheterocyclic group including one or more heteroatoms selected from N, Oand S, optionally substituted by one or more groups selected fromhalogen, C₁-C₆ alkyl and C(O)C₁-C₆ alkyl; wherein the alkyl groups areoptionally substituted by one or more halogen atoms, hydroxyl C₁-C₄alkoxy, C(O)NH₂, C(O)NHC₁-C₆ alkyl or C(O)N(C₁-C₆ alkyl)₂; orR²² and R²³ together with the nitrogen to which they are attached form a5- to 7-membered heterocycloalkyl group, the heterocycloalkyl groupincluding one or more further heteroatoms selected from N, O and S, theheterocycloalkyl group being optionally substituted by one or more Zgroups.

Embodiment 11

A compound according to any of embodiments 1 to 9 wherein R¹³ is (C₁-C₃alkyl)-NR²³C(O)R²², and wherein R²² and R²³ together with the atoms towhich they are attached form an oxo substituted 5- to 7-memberedheterocycloalkyl group, the heterocycloalkyl group including one or morefurther heteroatoms selected from N, O and S, the heterocycloalkyl groupbeing optionally further substituted by one or more Z groups.

Embodiment 12

A compound according to any preceding claim which is a compound ofFormula Ia

or a pharmaceutically acceptable salt or solvate thereof wherein R¹⁰ isas defined in any preceding claim.

Embodiment 13

A compound according to any preceding embodiment which is a compound ofFormula Ib

or a pharmaceutically acceptable salt thereof wherein B, R^(11a),R^(12a), R^(11b), R^(12b) and R¹³ are as defined anywhere above inrespect of a compound of Formula I.

Embodiment 14

A compound according to embodiments 1 to 12 which is a compound ofFormula Ic

or a pharmaceutically acceptable salt or solvate thereof wherein B,R^(11a), R^(12a), R^(11b), R^(12b), R¹³ and R¹⁵ and R¹⁷ are as definedanywhere above in respect of a compound of Formula

Embodiment 15

A compound according to embodiments 1 to 12 which is a compound ofFormula I

or a pharmaceutically acceptable salt or solvate thereof wherein B,R^(11a), R^(12a), and R¹³ are as defined anywhere above in respect of acompound of Formula I.

Embodiment 16

A compound according to embodiment 1 selected from:

-   3-(3-{2-[(E)-3,5-Diamino-6-chloro-pyrazine-2-carbonylimino]-1,3,8-triaza-spiro[4.5]decane-8-carbonyl}-benzenesulfonylamino)-propionic    acid dipropylcarbamoylmethyl ester;-   [4-(3-{2-[(Z)-3,5-Diamino-6-chloro-pyrazine-2-carbonylimino]-1,3,8-triaza-spiro[4.5]dec-8-yl}-3-oxo-propyl)-phenoxy]-acetic    acid [(2-hydroxy-ethyl)-methyl-carbamoyl]-methyl ester;-   [4-(3-{2-[(E)-3,5-Diamino-6-chloro-pyrazine-2-carbonylimino]-1,3,8-triaza-spiro[4.5]dec-8-yl}-3-oxo-propyl)-phenoxy]-acetic    acid cyclohexyl oxycarbonylmethyl ester;-   3-(3-{2-[(E)-3,5-Diamino-6-chloro-pyrazine-2-carbonylimino]-1,3,8-triaza-spiro[4.5]decane-8-carbonyl}-benzene    sulfonylamino)-propionic acid cyclohexyloxy carbonylmethyl ester;-   [4-(3-{2-[(E)-3,5-Diamino-6-chloro-pyrazine-2-carbonylimino]-1,3,8-triaza-spiro[4.5]dec-8-yl}-3-oxo-propyl)-phenoxy]-acetic    acid dimethylcarbamoylmethyl ester;-   [4-(3-{2-[(E)-3,5-Diamino-6-chloro-pyrazine-2-carbonylimino]-1,3,8-triaza-spiro[4.5]dec-8-yl}-3-oxo-propyl)-phenoxy]-acetic    acid dipropylcarbamoylmethyl ester;-   [4-(3-{2-[(E)-3,5-Diamino-6-chloro-pyrazine-2-carbonylimino]-1,3,8-triaza-spiro[4.5]dec-8-yl}-3-oxo-propyl)-phenoxy]-acetic    acid tert-butoxycarbonylmethyl ester;-   [4-(3-{2-[(E)-3,5-Diamino-6-chloro-pyrazine-2-carbonyl    imino]-1,3,8-triaza-spiro[4.5]dec-8-yl}-3-oxo-propyl)-phenoxy]-acetic    acid benzyloxycarbonylmethyl ester;-   [4-(3-{2-[(E)-3,5-Diamino-6-chloro-pyrazine-2-carbonylimino]-1,3,8-triaza-spiro[4.5]dec-8-yl}-3-oxo-propyl)-phenoxy]-acetic    acid diethylcarbamoylmethyl ester;-   [4-(3-{2-[(E)-3,5-Diamino-6-chloro-pyrazine-2-carbonylimino]-1,3,8-triaza-spiro[4.5]dec-8-yl}-3-oxo-propyl)-phenoxy]-acetic    acid 2-oxo-2-piperidin-1-yl-ethyl ester;-   [2-Chloro-4-(3-{2-[(E)-3,5-diamino-6-chloro-pyrazine-2-carbonylimino]-1,3,8-triaza-spiro[4.5]dec-8-yl}-3-oxo-propyl)-phenoxy]-acetic    acid dipropylcarbamoylmethyl ester;-   3-(3-{2-[(E)-3,5-Diamino-6-chloro-pyrazine-2-carbonylimino]-1,3,8-triaza-spiro[4.5]decane-8-carbonyl}-benzenesulfonylamino)-propionic    acid 2-oxo-2-(2-trifluoromethyl-pyrrolidin-1-yl)-ethyl ester;-   [2-Chloro-4-(3-{2-[(E)-3,5-diamino-6-chloro-pyrazine-2-carbonylimino]-1,3,8-triaza-spiro[4.5]dec-8-yl}-3-oxo-propyl)-phenoxy]-acetic    acid 2-(2-oxo-piperidin-1-yl)-ethyl ester;-   [4-(3-{2-[(E)-3,5-Diamino-6-chloro-pyrazine-2-carbonylimino]-1,3,8-triaza-spiro[4.5]dec-8-yl}-3-oxo-propyl)-phenoxy]-acetic    acid 2-morpholin-4-yl-2-oxo-ethyl ester;-   1-[(3-{2-[(E)-3,5-Diamino-6-chloro-pyrazine-2-carbonylimino]-1,3,8-triaza-spiro[4.5]decane-8-carbonyl}-benzenesulfonylamino)-methyl]-cyclobutanecarboxylic    acid dipropylcarbamoylmethyl ester;-   3-[3-(3-{2-[(E)-3,5-Diamino-6-chloro-pyrazine-2-carbonylimino]-1,3,8-triaza-spiro[4.5]decane-8-carbonyl}-phenyl)-ureido]-propionic    acid dipropylcarbamoylmethyl ester; and-   1-[(3-{2-[(E)-3,5-Diamino-6-chloro-pyrazine-2-carbonylimino]-1,3,8-triaza-spiro[4.5]decane-8-carbonyl}-benzenesulfonylamino)-methyl]-cyclobutanecarboxylic    acid-   2-oxo-2-(2-trifluoromethyl-pyrrolidin-1-yl)-ethyl ester;    or a pharmaceutically acceptable salt or solvate thereof.

Embodiment 17

A compound according to embodiment 16 selected from:

-   3-(3-{2-[(E)-3,5-Diamino-6-chloro-pyrazine-2-carbonylimino]-1,3,8-triaza-spiro[4.5]decane-8-carbonyl}-benzenesulfonylamino)-propionic    acid dipropylcarbamoylmethyl ester;-   [4-(3-{2-[(E)-3,5-Diamino-6-chloro-pyrazine-2-carbonylimino]-1,3,8-triaza-spiro[4.5]dec-8-yl}-3-oxo-propyl)-phenoxy]-acetic    acid dipropylcarbamoylmethyl ester; and-   [2-Chloro-4-(3-{2-[(E)-3,5-diamino-6-chloro-pyrazine-2-carbonylimino]-1,3,8-triaza-spiro[4.5]dec-8-yl}-3-oxo-propyl)-phenoxy]-acetic    acid dipropylcarbamoylmethyl ester;    or a pharmaceutically acceptable salt or solvate thereof.

Embodiment 18

A pharmaceutical composition, comprising:

-   -   a compound according to any of embodiments 1 to 16 and    -   one or more pharmaceutically acceptable excipients, diluents        and/or carriers.

Embodiment 19

A pharmaceutical composition according to embodiment 18, comprising:

-   -   one or more other therapeutic agents.

Embodiment 20

A pharmaceutical composition according to claim 15, comprising a secondagent, wherein the second agent is a CFTR potentiator of formula

Embodiment 21

A compound according to embodiments 1 to 17 for use in treating orpreventing a disease or condition mediated by blockade of the epithelialsodium channel.

Embodiment 22

A compound according to embodiment 21, wherein the disease or conditionis cystic fibrosis or COPD.

Embodiment 23

A compound according to embodiment 22, wherein the disease or conditionis cystic fibrosis.

Embodiment 24

Use of a compound according to any of embodiments 1 to 17 in themanufacture of a medicament for the prevention or treatment of a diseaseor condition mediated by blockade of the epithelial sodium channel

Embodiment 25

A method for preventing or treating a disease or condition mediated byblockade of the epithelial sodium channel in which an effective amountof a compound according to any of embodiments 1 to 17 is administered toa patient in need of such treatment.

Embodiment 26

A process for preparing a compound of formula I or a pharmaceuticallyacceptable salt or solvate thereof comprising the step of:

-   -   (a). reacting a compound of formula II

wherein R¹, R², R³, R⁴, R⁵, R⁶, R⁷, R⁸ and R⁹ are as defined in any ofembodiments 1 to 9, with a compound of formula III under conventionreaction conditions for acid-amine coupling

wherein R¹, R², R³, R⁴, R⁵, R⁶, R⁷, R⁸, R⁹ and R¹⁰ are as defined in anyof embodiments 1 to 9; or

-   -   (b) reacting a compound of formula IV

with a compound R¹³-L of formula V under convention reaction conditionswherein R¹, R², R³, R⁴, R⁵, R⁶, R⁷, R⁸, R⁹ and R¹³ are as defined in anyof embodiments;R^(10a) is(C₀-C₃alkylene)-B—X—(CR^(11a)R^(12a))_(m)—(CR^(11b)R^(12b))_(n)—(CR^(11c)R^(12c))_(p)—C(O)OHwherein B, X, R^(11a), R^(11b), R^(11c), R^(12a), R^(112b), R^(12c), m,n, and p are as defined in any of claims 1 to 9; andL is a leaving group.

1-12. (canceled)
 13. A method for treating chronic bronchitis comprisingadministering an effective amount of a compound chosen from:

or a pharmaceutically acceptable salt thereof, to a subject in need ofsuch treatment.
 14. The method according to claim 13 wherein thecompound is

or a pharmaceutically acceptable salt thereof.
 15. The method accordingto claim 14 wherein the compound is a pharmaceutically acceptable salt.16. The method of claim 15 wherein the pharmaceutically acceptable saltis a succinate salt.
 17. The method according to claim 13 wherein thecompound is

or a pharmaceutically acceptable salt thereof.
 18. The method accordingto claim 17 wherein the compound is a pharmaceutically acceptable salt.